Process for producing threo-3-amino-2-hydroxybutanoyl-aminoacetic acids, as well as novel intermediates therefor and process for producing them

ABSTRACT

Threo-3-protected amino-2-hydroxy-4-oxobutanoic acid or its esters represented by the formula:   &lt;IMAGE&gt;    wherein R1 represents naphthyl or a group of the formula   &lt;IMAGE&gt;  in which R6 and R7 represent individually hydrogen, halogen, amino, an amino protected with acyl, lower alkyloxycarbonyl or carbamoyl, hydroxy or protected with acyl, lower alkyloxycarbonyl or carbamoyl, lower alkoxy, lower alkyl or phenyl group; R2 represents an amine protected with acyl, lower alkyloxycarbonyl or carbamoyl; and R3 represents hydrogen, lower alkyl having 1 to 6 carbon atoms or benzyl.

This application is a divisional of application Ser. No. 96,693 filedNov. 23, 1979, now U.S. Pat. No. 4,281,180.

BACKGROUND OF THE INVENTION

Most of threo-3-amino-2-hydroxybutanoylaminoacetic acids preparedaccording to this invention are known from U.S. Pat. Nos. 4,029,547 and4,052,449, British Pat. Nos. 1,510,477 and 1,540,019.

For example, (2S,3R)-3-amino-2-hydroxy-4-phenylbutanoylaminoacetic acid(herein after referred to simply as bestatin) is a least toxic substanceisolated from culture filtrates of Streptomyces olivoreticuli as anaminopeptidase B inhibitor (referred to U.S. Pat. No. 4,029,547) by H.Umezawa, et al., which has been found to exhibit an increasing effect toimmuno-response such as delayed hypersthesia, activate in vivo defensemechanism and has inhibitive effects for cancer (referred to JapanesePatent Laid-Open Publication No. 117435/1977) and the compound isexpected to be useful as a pharmaceutical.

It is also known that several 3-amino-2-hydroxybutanoylaminoacetic acidsother than bestatin have equal or more inhibitory activity toaminopeptidase B than bestatin and a synthetic process therefor isdisclosed in British Pat. No. 1,510,477. In the process described in theBritish Patent, however, while (2S,3R)-3-amino-2-hydroxybutanoic acid,for example, as an intermediate for bestatin is synthesized through thesteps shown below, the process has various defects as detailed later.##STR3##

(R)-phenylalanine (1) is converted to a benzyloxycarbonyl derivative(2), which is then condensed with 3,5-dimethylpyrazole by usingdicyclohexylcarbodiimide. The 3,5-dimethylpyrazolide (3) is reduced tobenzyloxycarbonyl-(R)-phenylalaninal (4) with lithium aluminum hydride,which is then changed to the corresponding adduct (5) by reaction withsodium hydrogen sulfite and further to the cyanohydrin (6) throughreaction with a cyanide. The above derivative is hydrolyzed under anacidic condition into (2RS,3R)-3-amino-2-hydroxy-4-phenylbutanoic acid(7), which is again allowed to react with a benzyloxycarbonylatingreagent to convert to(2RS,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-butanoic acid (8). Theacid is fractionally crystallized into(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid (9) byusing brucine.

Although the above process can be conducted with no troubles in alaboratory scale, it is not suited to large scale production so that theprocess includes the use of much expensive (R)-phenylalanine which isnot a natural amino acid, the use of highly ignitable lithium aluminumhydride for the reduction of the pyrazolide, as well as the use of avery poisonous cyanide in synthesis of cyanohydrin.

In view of the above, the inventors have made an earnest study seekingfor a process suited to mass production with no foregoing defects and,accomplished this invention.

The process according to this invention can produce the end products ata high yield without using an expensive amino acid such as(R)-phenylalanine and with no use of a cyanide, and thus it is muchsuited to the mass production.

SUMMARY OF THE INVENTION

This invention concerns a process for producingthreo-3-amino-2-hydroxybutanoylaminoacetic acids and esters thereofwhich comprises the steps of:

(A) reacting a N-protected 2-oxoethylamine represented by the generalformula: ##STR4## fractionally crystallized into(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid (9) byusing brucine.

Although the above process can be conducted with no troubles in alaboratory scale, it is not suited to large scale production so that theprocess includes the use of much expensive (R)-phenylalanine which isnot a natural amino acid, the use of highly ignitable lithium aluminumhydride for the reduction of the pyrazolide, as well as the use of avery poisonous cyanide in synthesis of cyanohydrin.

In view of the above, the inventors have made an earnest study seekingfor a process suited to mass production with no foregoing defects and,accomplished this invention.

The process according to this invention can produce the end products ata high yield without using an expensive amino acid such as(R)-phenylalanine and with no use of a cyanide, and thus it is muchsuited to the mass production.

SUMMARY OF THE INVENTION

This invention concerns a process for producingthreo-3-amino-2-hydroxybutanoylaminoacetic acids and esters thereofwhich comprises the steps of:

(A) reacting a N-protected 2-oxoethylamine represented by the generalformula: ##STR5## wherein R₁ represents a naphthyl or a group of theformula: ##STR6## in which R₆ and R₇ individually represent hydrogen,halogen, amino or a protected amino, hydroxy or a protected hydroxy, alower alkoxy, a lower alkyl or phenyl and R₂ represents a protectedamino, with glyoxylic acid or its ester represented by the generalformula: ##STR7## wherein R₃ represents hydrogen or an ester residue, tochange into threo-3-protected amino-2-hydroxy-4-oxobutanoic acid or itsester represented by the general formula: ##STR8## wherein R₁, R₂ and R₃have the same meaning as above; (B) reducing the above compound intothreo-3-protected amino-2-hydroxybutanoic acid or its ester representedby the general formula: ##STR9## wherein R₁, R₂ and R₃ have the samemeanings as above; (C) subjecting the above compound, as may berequired, to step(s) of (a) ester residue elimination, (b) opticalresolution and/or (c) amino protecting group removal to obtainthreo-3-amino-2-hydroxybutanoic acid represented by the general formula:##STR10## wherein R₁ has the same meaning as above and R₂ ' representsamino or a protected amino, and

(D) condensing the above compound in a conventional manner for theformation of a peptide coupling with an aminoacetic acid represented bythe general formula: ##STR11## wherein R₄ represents an alkyl having 3-4carbon atom number or 3-guanidinopropyl, after protecting, as required,functional group therein not relevant to the reaction and then removingthe protecting group for the functional group to thereby prepare3-amino-2-hydroxy-butanoylaminoacetic acid represented by the generalformula: ##STR12## wherein R₁ and R₄ have the same meanings as above.

This invention also concerns threo-3-amino-2-hydroxy-4-oxobutanoic acidsof the general formula (III) and esters thereof, a process for theproduction thereof, as well as a process for producing threo-3-protectedamino-2-hydroxybutanoic acids of the general formula (IV).

DETAILED DESCRIPTION OF THE INVENTION

In the starting material of the general formula (I) in this invention,R₁ may be a naphthyl, for example, 1-naphthyl and 2-naphthyl or may be agroup of the formula ##STR13## in which R₆ and R₇ may be identical ordifferent to each other.

R₆ and R₇ may be halogen such as chlorine, bromine and fluorine, a loweralkoxy having 1-6 carbon atom, for example, methoxy, ethoxy, ethoxy,propoxy, butoxy, pentyloxy and hexyloxy and a lower alkyl having 1-6carbon atom, for example methyl, ethyl, propyl, butyl, penthyl andhexyl. The lower alkoxy or lower alkyl may be branched.

The group represented by the formula ##STR14## includes, for example,phenyl, chlorophenyl, dichlorophenyl, fluorophenyl, aminophenyl,hydroxyphenyl, methoxyphenyl, ethoxyphenyl, n-propoxyphenyl,iso-propoxyphenyl, n-butoxyphenyl, iso-butoxyphenyl, sec-butoxyphenyl,methylphenyl, n-propylphenyl, iso-propylphenyl, n-butylphenyl,iso-butylphenyl, sec-butylphenyl, biphenyl, dihydroxyphenyl,dimethoxyphenyl and hydroxymethoxyphenyl. Substituents on the benzenenuclei may take any possible positions, that is, o-, m- or p-position;o- and m-positions, p- and o-positions or p- and m-positions.

The protecting group on the protected amino in R₂ includes an acyl, forexample, formyl, a lower alkylcarbonyl which may have substituents suchas halogen with no interference to the reaction, for example, acetyl,chloroacetyl, dichloroacetyl, bromoacetyl, 2-chloropropionyl and2-bromopropionyl or benzoyl which may have substituents such as a loweralkoxy, a lower alkyl or halogen on the phenyl ring not interferringwith the reaction; a lower alkyloxycarbonyl which may have substituentssuch as halogen not interferring with the reaction, for example,methoxycarbonyl, ethoxycarbonyl, iso-propyloxycarbonyl,iso-butyloxycarbonyl, t-butyloxycarbonyl, t-amyloxycarbonyl and2,2,2-trichloroethoxycarbonyl, a cycloalkyloxycarbonyl, for example,cyclopentyloxycarbonyl and cyclohexyloxycarbonyl; a benzyloxycarbonylwhich may have substituents such as a lower alkoxy, a lower alkyl andhalogen on the phenyl ring not interferring with the reaction, forexample, benzyl oxycarbonyl, chlorobenzyloxycarbonyl,nitrobenzyloxycarbonyl, methoxybenzyloxycarbonyl,methylbenzyloxycarbonyl; carbamoyl, a lower alkylcarbamoyl, for example,methylcarbamoyl, ethylcarbamoyl, propylcarbamoyl and butylcarbamoyl andphenylcarbamoyl, as well as phthalyl. The alkyl and phenyl in thephthalyl or carbamoyl may have substituents not interferring with thereaction.

The above protecting groups can also be used as the protecting groupsfor R₆ and R₇ if they are amino or hydroxy groups.

A N-protected 2-oxoethylamine of the general formula (I) employed as astarting material in this invention includes many novel compounds, andthey can be synthesized, in the same manner as that for known compoundsfrom a methylketon represented by the general formula:

    R.sub.1 -COCH.sub.3                                        (VIII),

wherein R₁ has the same meaning as above, by brominating it into acorresponding bromomethylketone, reacting the brominated ketone withhexamethylenetetramine, hydrolyzing it into a corresponding2-oxoethylamine and then protecting the amino groups therein throughreaction with an appropriate acylating agent.

The compound of the general formula (I) includes, sepecifically,

N-(2-oxo-2-phenylethyl)acetamide,

N-(2-oxo-2-phenylethyl)benzamide,

N-(2-oxo-2-phenylethyl)phthalimide,

2-methoxycarbonylaminoacetophenone,

2-t-butyloxycarbonylaminoacetophenone,

N-[2-oxo-2-(4-hydroxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-hydroxyphenyl)ethyl]chloroacetamide,

2-ethoxycarbonylamino-4'-hyroxyacetophenone,

N-[2-oxo-2-(3-hydroxyphenyl)ethyl]benzamide,

2-t-butyloxycarbonylamino-3'-hydroxyacetophenone,

N-(2-oxo-2-phenylethyl)chloroacetamide,

2-t-butyloxycarbonylaminoacetophenone,

N-[2-oxo-2-(4-chlorophenyl)ethyl]acetamide,

N-[2-oxo-2-(3-chlorophenyl)ethyl]acetamide,

N-[2-oxo-2-(4-methylphenyl)ethyl]acetamide,

N-[2-oxo-2-(3-methylphenyl)ethyl]acetamide,

N-[2-oxo-2-(1-naphthyl)ethyl]acetamide,

N-[2-oxo-2-(2-naphthyl)ethyl]acetamide,

N-[2-oxo-2-(4-fluorophenyl)ethyl]acetamide,

N-[2-oxo-2-(4-bromophenyl)ethyl]acetamide,

N-[2-oxo-2-(2-methoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(3-methoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-methoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(2-ethoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(3-ethoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-ethoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-n-propoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-iso-propoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-n-butoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-isobutoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-sec-butoxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(2-ethylphenyl)ethyl]acetamide,

N-[2-oxo-2-(3-ethylphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-ethylphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-n-propylphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-iso-propylphenyl)ethyl]benzamide,

N-[2-oxo-2-(4-n-butylphenyl)ethyl]acetamide,

N-[2-oxo-2-(4-iso-butylphenyl)ethyl]benzamide,

N-[2-oxo-2-(4-sec-butylphenyl)ethyl]acetamide,

N-[2-oxo-2-(2-hydroxyphenyl)ethyl]acetamide,

N-[2-oxo-2-(3-hydroxyphenyl)ethyl]acetamide, and

N-[2-oxo-2-(3,4-dihydroxyphenyl)ethyl]acetamide.

The ester residues of R₃ in the compound represented by the generalformula (II) have no particular restrictions so long as they result inno interference with the reaction and those employed usually include alower alkyl having 1-6 carbon atom number such as methyl, ethyl, propyl,butyl, pentyl and hexyl or benzyl, in which these lower alkyl or benzylgroup may have substituents such as halogen not interferring with thereaction.

The compound of the general formula (II) may include, for example,glyoxylic acid, its methyl, ethyl and benzyl esters.

The reaction of N-protected 2-oxoethylamide of the general formula (I)and glyoxylic acid or its ester of the general formula (II) in thisinvention is usually carried out in water, an organic solvent or a mixedsolvent of water and an organic solvent in the presence of a base.

Organic solvents employed as the mixed solvent with water have noparticular restriction so long as they are watermiscible and they,preferably, include polar solvents such as lower alcohols, for example,methanol, ethanol and propanol; ketones, for example, acetone andmethylethylketone; acetonitrile, tetrahydrofuran, dioxane,dimethylformamide, dimethylacetamide, dimethylsulfoxide.

Organic solvents in which the reaction is conducted also have noparticular restriction so long as they can dissolve the startingmaterials and they, preferably, include in addition to the solventsreferred to aboe, esters, for example, a lower alkylester of acetic acidsuch as methyl acetate and ethyl acetate; ethers, for example,diethylether and isopropylether, and halogenated hydrocarbons, forexample, chloroform and carbontetrachloride.

The bases used herein include inorganic bases such as hydroxides,carbonates and hydrogen carbonates of alkali or alkaline earth metals,for example, potassium hydroxide, sodium hydroxide, calcium hydroxide,potassium carbonate, sodium carbonate, potassium hydrogen carbonate andsodium hydrogen carbonate, and ammonia; organic bases such as alkalimetal salts of organic acids, for example, alkali metal salts of loweralkyl carboxylic acids such as potassium acetate, sodium acetate, sodiumformate and potassium propionate, aromatic amines, for example, pyridineand aliphatic amines, for example, trimethylamine and triethylamine. Forthe reaction in water or a watercontaining organic solvent, inorganicbases, particularly, hydrogen carbonates of alkali metals, for example,sodium hydrogen carbonate and potassium hydrogen carbonate arepreferred. For the reaction in an organic solvent, are used organicbases, preferably, alkylamines and, in particular, triethylamine.

The amount of the bases employed has no particular restriction so longas the reaction solution is kept between slightly acidic and alkaline,and it is preferred to use them in an amount, for example, 0.5-10 mol,and usually, 1-2 mol per mol of glyoxylic acid or its ester representedby the general formula (II).

The reaction can be carried out at a reaction temperature from 0° C. tothe boiling point of a solvent and, usually, from room temperature up to60° C.

While the reaction time varies depending on the reaction temperature,starting compounds and the solvent, it is not determined specificallybut the reaction is completed in about 2-100 hours and, usually, inabout 12 -20 hours at the reaction temperature from room temperature upto 60° C.

The glyoxylic acid or its ester represented by the general formula (II)used in this invention has not particular restriction in its amount andit is employed, usually, in a wide range between 0.2-10 mol andpreferably, 1-2 mol per mol of N-protected 2-oxoethylamine representedby the general formula (I).

For the glyoxylic acid of the general formula (II) in which R₃represents hydrogen atom, an inexpensive aqueous solution can be usedwith no troubles for the reaction.

Through the reaction of the compound of the general formula (I) and thecompound of the general formula (II) in this way, the compound of thegeneral formula (III) can be formed. The compound formed is isolatedfrom the reaction mixture, for example, as follows.

(a) For the compound in which R₃ is hydrogen atom

(1) If water is used for the reaction solvent, the reaction mixture isacidified with an acid to precipitate crystals, which are separated byfiltration.

(2) If a mixed solvent of water and an organic solvent is used for thereaction solvent, the organic solvent is at first removed under reducedpressure and then the residue is made acidic by the addition of an acidto precipitate crystals, which are separated by filtration.

(b) For the compound in which R₃ is an ester residue

Since an organic solvent or a water-containing solvent is usuallyemployed as the reaction solvent in this case, the reaction mixture isconcentrated under reduced pressure after the reaction to remove theorganic solvent. Then, water is added to the residue, which is thenextracted with an organic solvent such as ethyl acetate. The ethylacetate phase is washed with water and dried over a drying agent such asanhydrous sodium sulfate. The solvent is distilled off under reducedpressure, the residue is triturated with n-hexane and then precipitatedcrystals are collected by filtration.

All of the threo-(2RS)-3-amino-2-hydroxy-4-oxobutanoic acids or theesters thereof represented by the general formula (III) obtained arenovel compounds and they include as typical examples:

threo-(2RS)-3-acetylamine-2-hydroxy-4-oxo-4-phenylbutanoic acid,

ethyl ester ofthreo-(2RS)-3-acetylamine-2-hydroxy-4-oxo-4-phenylbutanoic acid,

threo-(2RS)-3-chloroacetylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid,

methyl ester ofthreo-(2RS)-3-chloroacetylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid,

threo-(2RS)-3-methoxycarbonylamino-2-hydroxy-4-oxo-4-phenylbutanoicacid,

threo-(2RS)-3-t-butoxycarbonylamino-2-hydroxy-4-oxo-4-phenylbutanoicacid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-hydroxyphenyl) butanoicacid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(4-hydroxyphenyl) butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-hydroxyphenyl) butanoicacid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(3-hydroxyphenyl) butanoicacid,

threo-(2RS)-3-ethoxycarbonylamino-2-hydroxy-4-oxo-4-(4-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-t-butoxycarbonylamino-2-hydroxy-4-oxo-4-(3-hydroxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(1-naphthyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-naphthyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-fluorophenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-bromophenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-methoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-methoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid,

methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-ethoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-ethoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-ethoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-n-propoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-iso-propoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-n-butoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-iso-butoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-sec-butoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-ethylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-ethylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-ethylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-n-propylphenyl)butanoicacid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(4-iso-propylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-n-butylphenyl)butanoicacid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(4-iso-butylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-sec-butylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-hydroxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-hydroxyphenyl)butanoicacid, and

threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3,4-dihydroxyphenyl)butanoicacid.

The threo-(2RS)-3-protected amino-2-hydroxy-4-oxobutanoic acid or itsester obtained and represented by the general formula (III) can beconverted with ease to threo-(2RS)-3-amino-2-hydroxybutanoic acid or itsester represented by the general formula (IV) by reduction.

The reduction process used herein can be selected with no particularrestriction provided that it can reduce the carbonyl group directlycombined to an aromatic nuclei to methylene group and the processincludes, for example, catalytic hydrogenation using palladium such aspalladium black, palladium carbon and palladium barium sulfate or Raneynickel, reduction with lithium aluminum hydride, sodiumbis(2-methoxyethoxy) aluminum hydride in the presence of anhydrousaluminum chloride, reduction with an acidic zinc or tin, and reductionwith metal sodium or metal lithium in liquid ammonia. The catalytichydrogenation using palladium such as palladium black and palladiumcarbon or Raney nickel is particularly preferred.

The reduction process using palladium or Raney nickel is carried out bydissolving or suspending threo-3-protected amino-2-hydroxy-4-oxobutanoicacid or its ester represented by the general formula (III) into asolvent and adding palladium or Raney nickel thereto, and reducing in ahydrogen atmosphere. The hydrogen may be used at an atmospheric pressureor an elevated pressure in an autoclave, and the reduction can beproceeded rapidly under an elevated pressure.

While the reaction solvent is not particularly restricted so long as itcan dissolve threo-3-protected amino-2-hydroxy-4-oxobutanoic acid or itsester represented by the general formula (III) even in a small amount,the use of lower alcohols such as methanol ethanol and propanol, loweralkylcarboxylic acids such as acetic acid and propionic acid, watercontaining them or mixed solvents containing them with other organicsolvents is preferred. While reaction temperature from 0° C. up to 150°C. can be used, the reaction is, preferably, carried out at 40°-100° C.in a solvent containing acetic acid or methanol.

3-protected amino-2-hydroxybutanoic acid or its ester formed andrepresented by the general formula (IV) can be isolated with ease fromthe reaction mixture by filtrating out the catalyst, then concentratingthe filtrate and adding an appropriate solvent to the residue toprecipitate crystals and collecting the precipitated crystals byfiltration.

Those compounds represented by the general formula (IV) in which R₂ isprotected by the protecting groups such as chloroacetyl, dichloroacetyl,t-butoxycarbonyl, t-pentyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl,cyclopentyloxycarbonyl, cyclohexyloxycarbonyl, and phthalyl areespecially preferred since, after optical resolution as required, theycan be allowed to directly react with aminoacetic acid of the generalformula (VI) to prepare threo-3-amino-2-hydroxybutanolacetic acidrepresented by the general formula (VII).

Threo-3-protected amino-2-hydroxybutanoic acid or its ester representedby the general formula (IV) includes as its typical examples:

threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid, ethyl esterof threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-phenylbutanoic acid,

threo-(2RS)-3-chloroacetylamino-2-hydroxy-4-phenylbutanoic acid, methylester of threo-(2RS)-3-chloroacetylamino-2-hydroxy-4-phenylbutanoicacid,

threo-(2RS)-3-methoxycarbonylamino-4-phenylbutanoic acid,

threo-(2-RS)-3-t-butoxycarbonylamino-4-phenylbutanoic acid,

threo-(2RS)-3-phthalimino-4-phenylbutanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-(4-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-(3-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-(2-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(1-naphthyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-naphthyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-fluorophenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-bromophenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-methoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-methoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid,

methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-ethoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-ethoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-ethoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-n-propoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-iso-propoxyphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-n-butoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-iso-butoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-sec-butoxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-ethylphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-ethylphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-ethylphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-n-propylphenyl)butanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-(4-iso-propylphenyl)butanoicacid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-n-butylphenyl)butanoic acid,

threo-(2RS)-3-benzoylamino-2-hydroxy-4-(4-isobutylphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-sec-butylphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-hydroxphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-hydroxyphenyl)butanoic acid,

threo-(2RS)-3-acetylamino-2-hydroxy-4-(3,4-dihydroxyphenyl)butanoicacid,

and (2S,3R) type optical isomers thereof, as well as lower alkyl esters,for example, methyl, ethyl, propyl and butyl esters; and benzyl andchlorobenzyl esters of these acids.

If the compounds represented by the general formula (IV) are esters,they can be converted to threo-3-amino-2-hydroxybutanoic acids of thegeneral formula (IV) by removing the ester residues in a conventionalmanner such as hydrolysis or reduction and further eliminating, ifrequired, the amino protecting groups.

A threo-(2RS) compound represented by the general formula (IV) may beoptically resolved to (2S,3R) and (2R,3S) isomers as required. In thecase of a compound of the general formula (IV), in which R₃ is hydrogen,(a) it is directly resolved with an optically active base such asbrucine and S(-)-or R(+)-1-phenylethylamine, (b) it is resolved with anoptically active sulfonic acid such as camphorsulfonic acids orbromocamphorsulfonic acids after an amino protecting group in thecompound is removed in the usual manner. In the case of a compound ofthe general formula (IV), in which R₃ is an ester residue, (c) it isresolved in the above-mentioned procedure after the ester residue isremove in the usual manner or (d) it is resolved with an opticallyactive acid such as tartaric acid derivatives and camphorsulfonic acidsafter an amino protecting group is removed by the general method.

The solvent used in the optical resolution is properly selecteddepending on the compound of the general formula (IV) and the type of aresolving reagent.

If a (2S,3R) isomer obtained has an ester residue, it can be convertedto (2S,3R)-3-amino-2-hydroxybutanoic acid represented by the generalformula (V) by elimination of the ester residue in a conventionalmanner.

Condensation of compound represented by the general formula (IV) or (V)and aminoacetic acid of the general formula (VI) is carried out in aconventional manner for the formation of peptide linkage whileprotecting those functional groups not relevant to the reaction asrequired.

3-amino-2-hydroxybutanoylaminoacetic acid represented by the generalformula (VII) which is the aimed compound of this invention can beprepared by removing the protecting groups for the functional groupsafter the end of the condensing reaction.

Protection for the functional groups not relevant to the reaction, forexample, the protections for the amino group of R₂ ' in the compound ofthe general formula (V) and for the carboxylic group in the compound ofthe general formula (VI) can be conducted in a conventional manner. Theprotection for the amino group, for example, with benzyloxycarbonylgroup can be conducted by reaction of the compound of the generalformula (V) in which R₂ ' is an amino group and benzyloxycarbonylchloride in Schotten-Baumann procedure, or reaction withbenzyloxycarbonylating agent such as p-nitrophenyl benzyloxycarbonate,benzyloxycarbonylazide, benzyloxycarbonyl N-hydroxysuccinimide ester,benzyl S-4,6-dimethylpyrimide-2-yl-thiocarbonate, in the presence of atertially organic base such as triethylamine or N-methylmorpholine in anaqueous organic solvent, for example, dioxane, tetrahydrofurane,acetonitril or dimethylformamide.

The condensing methods between the acid of the general formula (V) andthe aminoacetic acid of the general formula (VI) include, for example, acarbodiimide process using dicyclohexylcarbodiimide and1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, an azide process, amixed acid anhydride process using ethyl chloroformate and isobutylchloroformate, an active ester process using cyanomethyl ester, vinylester, substituted and non-substituted phenyl ester, thiophenyl esterand hydroxysuccinimide ester, an O-acylhydroxylamine derivative processusing acetoxime and cyclohexanone oxime, an N-acyl compound processusing carbodiimidazole.

Solvents employed in formation of the usual peptide linkage may be alsoused as the solvents for the condensation in this invention. Thefollowing solvents can, for example, be used; ethers such asdiethylether, tetrahydrofuran and dioxane, esters such as ethyl acetateand methyl acetate, ketones such as acetone and methylethylketone,halogenated hydrocarbons such as methylene chloride and chloroform,amides such as dimethylformamide and dimethylacetamide and nitriles suchas acetonitrile.

After the end of the condensing reaction, protecting groups are removedin the usual manner for the elimination of the protecting groups in thepeptide chemistry such as catalytic reduction using palladium as acatalyst, acidolysis with hydrogen bromide in acetic acid,trifluoroacetic acid, hydrogen fluoride, hydrogen chloride in an organicsolvent, saponification with an alkali, reducting with metal sodium inliquid ammonia.

The representative examples of the finally aimed products represented bythe general formula (VII) are as follows:

threo-(2RS)-3-amino-2-hydroxy-4-(1-naphthyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(2-naphthyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-fluorophenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(2-ethoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(3-ethoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-ethoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-n-propoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-propoxyphenyl)butanoyl(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-n-butoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-butoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-sec-butoxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(2-ethylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(3-ethylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-ethylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-n-propylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-propylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-n-butylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-butylphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-sec-butylphenyl)butanyol-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(2-hydroxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(3-hydroxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(3,4-dihydroxyphenyl)butanoyl-(S)-leucine,

threo-(2RS)-3-amino-2-hydroxy-4-(4-chlorophenyl)butanyol-(S)-leucine,and

threo-(2RS)-3-amino-2-hydroxy-4-(3-chlorophenyl)butanoyl-(S)-leucine,

or those compounds in which threo-(2RS) configuration is replaced with(2S, 3R) form and (S)-leucine is replaced with (RS)- or (R)-leucine,(S)-, (RS)- or (R)-valine, (S)-, (RS)- or (R)-norvaline, (S)-, (RS)- or(R)-isoleucine, (S)-, (RS)- or (R)-norleucine, (S)-, (RS)- or(R)-tert-leucine or (S)-, (RS)- or (R)-arginine, especially

(2S,3R)-3-amino-2-hydroxy-4-(4-aminophenyl)butanoyl-(S)-leucine,

(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-leucine(bestatin),

(2S,3R)-3-amino-2-hydroxy-4-(2-hydroxyphenyl)butanoyl-(S)-leucine(o-hydroxybestatin),

(2S,3R)-3-amino-2-hydroxy-4-(3-hydroxyphenyl)butanoyl-(S)-leucine(m-hydroxybestatin),

(2S,3R)-3-amino-2-hydroxy-4-(4-hydroxyphenyl)butanoyl-(S)-leucine(p-hydroxybestatin),

(2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucine,

(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-valine,

(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-norvaline,

(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-arginine, and

(2S,3R)-3-amino-2-hydroxy-4-(4-hydroxphenyl)butanoyl-(S)-arginine,

as well as physiologically non-toxic salts thereof, for example, saltsof hydrochloric acid and acetic acid.

The above-mentioned compounds include many novel compounds together withseveral known compounds. As these novel compounds have inhibitory effectagainst aminopeptidase B, inhibit the formation of bradykinin andexhibit anti-inflammatory effects as the known compounds, they areexpected to be useful as pharmaceutical medicine for various deceases.

Among the compounds of the general formula (VII) which are preparedaccording to this invention, those compound are novel wherein R₁represents a naphthyl or a group of the formula ##STR15## in which R₇represents hydrogen and R₆ represents fluorine, lower alkyl (exceptingmethyl), lower alkyloxy, phenyl, m- or o-hydroxy, or both of R₆ and R₇represent groups other than hydrogen, and the aminopeptidase Binhibitory activity of the representative compounds is as follows.

Inhibitory Activity to Aminopeptidase B Test Method

Measurement for the inhibitory activity to aminopeptisase B wasconducted by the method reported by Hopsu et al. while somewhatmodifying the process [V. K. Lopsu, K. K. Makinen, G. G. Glenner;Archives of Biochemistry and Biophysics, 114, 557 (1966)].

A mixed solution (pH 7.0) prepared by adding 1.0 ml of a 0.1 Mtris-hydrochloric acid buffer solution and 0.7 ml of a solutioncontaining a test specimen to 0.3 ml of 0.1 mM arginine-β-naphthylamidewas heated at 37° C. for 4 minutes. Thereafter, 0.2 ml of anaminopeptidase B solution purified by the same enzyme purificationprocess as in the process by Hopsu et al. using Sephadex G-100(registered Trademark) was added and reacted at 37° C. for 30 minutes.Then, 0.6 ml of a 1.0 M acetic acid buffer solution (pH 4.2) containingGarnet GBC (o-aminoazotoluenediazonium salt) at 1.0 mg/ml concentrationand containing Tween 20 (registered trademark) at 1.0% concentration wasfurther added and left for 15 minutes at room temperature, after whichabsorption ratio (a) at 530 nm was measured. The absorption ratio (b)for the blind solution using only the buffer solution containing nospecimen was simultaneously measured, and the inhibitory ratio toaminopeptidase B was calculated as (b-a)/b×100.

Result

Inhibitory ratios for each of the specimens at several concentrationswere determined by the above test method, from which 50% inhibitoryratios (IC₅₀) were deduced. The results are shown in Table 1.

                                      TABLE 1                                     __________________________________________________________________________    No.                                                                              Compound                  IC.sub.50 (μg/ml)                             __________________________________________________________________________    1. (2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)                                                           0.006                                               butanoyl-(S)-leucine                                                       2. Threo-(2RS)-3-amino-2-hydroxy-4-(2-methoxyphenyl)                                                       2.9                                                 butanoyl-(S)-leucine                                                       3. Threo-(2RS)-3-amino-2-hydroxy-4-(3-methoxyphenyl)                                                       0.04                                                butanoyl-(S)-leucine                                                       4. Threo-(2RS)-3-amino-2-hydroxy-4-(1-naphthyl)                                                            1.5                                                 butanoyl-(S)-leucine                                                       5. Threo-(2RS)-3-amino-2-hydroxy-4-(2-naphthyl)                                                            0.10                                                butanoyl-(S)-leucine                                                       6. Threo-(2RS)-3-amino-2-hydroxy-4-(4-phenyl                                                               3.6                                                 phenyl)butanoyl-(S)-leucine                                                7. Threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-                                                                 0.12                                                propylphenyl)butanoyl-(S)-leucine                                          8. Threo-(2RS)-3-amino-2-hydroxy-4-(4-iso                                                                  2.8                                                 butylphenyl)butanoyl-(S)-leucine                                           9. Threo-(2RS)-3-amino-2-hydroxy-4-(3,4-                                                                   1.8                                                 dihydroxyphenyl)butanoyl-(S)-leucine                                       10.                                                                              Threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-                                                                 6.8                                                 propoxyphenyl)butanoyl-(S)-leucine                                            Threo-(2RS)-3-amino-2-hydroxy-4-(4-iso-                                                                 4.5                                                 butoxyphenyl)butanoyl-(S)-leucine                                             Threo-(2RS)-3-amino-2-hydroxy-4-(4-                                                                     0.03                                                fluorophenyl)butanoyl-(S)-leucine                                             Threo-(2RS)-3-amino-2-hydroxy-4-(2-naphthyl)                                                            0.03                                                butanoyl-(S)-isoleucine                                                       (2R,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)                                                           0.07                                                butanoyl-(S)-valine                                                           Threo-(2RS)-3-amino-2-hydroxy-4-(2-                                                                     0.53                                                hydroxyphenyl)butanoyl-(S)-leucine                                            Threo-(2RS)-3-amino-2-hydroxy-4-(3-                                                                     0.09                                                hydroxyphenyl)butanoyl-(S)-leucine                                         __________________________________________________________________________

This invention is to be described specifically by way of examples.

EXAMPLE 1 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid

N-[2-oxo-2-(4-methoxyphenyl)ethyl]acetamide (82.8 g, 0.400 mol), 95.8 g(1.14 mol) of sodium hydrogen carbonate and 66.3 g (0.720 mol) ofglyoxylic acid monohydrate were dissolved in a mixture of 700 ml ofmethanol and 200 ml of water. When the solution was allowed to react at50°-60° C., the reaction was complete after 4 hours. The reactionmixture was concentrated under reduced pressure to dryness. The residuewas dissolved in water and then washed with ethyl acetate. The aqueousphase was adjusted with hydrochloric acid to pH 1-2. Deposited crystalswere collected by filtration, washed with water and dried in vacuo.Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid was obtained. Yield 102.3 g (91.0%). Mp. 193°-195° C.(decompositon). NMR Spectrum (DMSO-d₆) δ=1.9 (s, 3H; CH₃ -CO), 3.8 (s, 3H; CH₃ -O), 4.5 (d,H,J=4Hz; CH-OH), 5.7 (dd, H; CH-NH), 7.1, 7.9(d,d,2H,2H,J=9Hz; ##STR16##

(2) Preparation ofthreo-(2RS)-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid (30.0 g, 0.107 mol) was dissolved in 300 ml of methanol and 1.50 gof 5% palladium carbon was added. When the solution was reduced in anautoclave at a temperature of 40° C. and at a hydrogen pressure of 25kg/cm², the reaction was completed after about 3 hours.

The catalyst was removed by filtration and the filtrate was concentratedunder reduced pressure. To the residue was added 100 ml of ethyl acetateand precipitated crystals were collected by filtration, washed withethyl acetate and dried in vacuo.Threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl) butanoic acidwas obtained. Yield 26.5 g (92.7%). Mp. 174°-176° C. NMR Spectrum (CF₃COOD), δ=2.3 (s, 3H; CH₃ -CO), 3.1 (d, 2H, J=8 Hz; CH₂), 4.0 (s, 3H; CH₃-O), 4.6 (d, H, J=2Hz; CH-OH), 7.0, 7.3 (d, d, 2H, 2H, J=9 Hz; ##STR17##

(3) Preparation of(2S,3R)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl) butanoic acid(10.0 g, 37.0 mmol) and 4.8 g (37.0 mmol) of S(-)-1-phenylethylaminewere dissolved under heating in 75 ml of ethanol. They were then allowedto cool at room temperature. Deposited crystals were collected byfiltration, washed with a small amount of ethanol and dried in vacuo.4.56 g of crystals were obtained. [α]_(D) ²⁰ +32.1° (c=1.9 methanol).

The crystals (4.50 g) were dissolved in 30 ml of ethanol under heatingand allowed to cool to room temperature. Precipitated crystals havecollected by filtration, washed with a small amount of ethanol anddried. The S(-)-1-phenylethylamine salt of(2S,3R)-3-acetylamino-2-hydroxy-(4-methoxyphenyl) butanoic acid wasobtained. Yield 4.24 g. Mp. 194°-195° C. [α]_(D) ²⁰ +32.8° (c=0.5,methanol).

(4) Preparation of(2S,3R)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid

The s(-)-phenylethylamine salt of(2S,3R)-2-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid (2.79g, 7.20 mmol) was added to 15 ml of 0.5 N sodium hydroxide aqueoussolution and S(-)-1-phenylethylamine was extracted by the addition ofeach 15 ml of ethyl acetate for three times.

The aqueous phase was separated and adjusted with N hydrochloric acid topH 1-2 and concentrated to dryness under reduced pressure. To theresidue was added 20 ml of acetone, insoluble compounds were removedhyfiltration and then the filtrate was concentrated to dryness underreduced pressure. Ethyl acetate (30 ml) was added to the residue.Deposited crystals were collected by filtration, washed with ethylacetate and dried anoic invacuo.(2S,3R)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid wasobtained. Yield 1.32 g (68.1% ). [α]_(D) ²² +27.7° (c=1.1, methanol).

(5) Preparation of (2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid (2S,3R)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid (4.89 g, 18.3 mmol) was added to a mixture of 22 ml of 2.5N hydrochloric acid and 22 ml of dioxane. When the solution was heatedat 60° C., the reaction was completed after 8 hours.

The reaction solution was concentrated under reduced pressure todryness. The residue was dissolved in water and again concentrated underreduced pressure to dryness. The residue was dissolved in 20 ml of waterand insoluble compounds were separated out by filtration. Then, theaqueous phase was adjusted with 2 N sodium hydroxide aqueous solution topH 5-6 and chilled in an ice bath. Precipitated crystals were collectedby filtration, washed with water and dried in vacuo.(2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid was obtained.Yield 3.28 g (79.6%). [α]_(D) ²⁰ +26.9° (c=1, N HCl). Mp. 230°-232° C.(decomposition). NMR Spectrum (CF₃ COOD) δ=3.2 (dd, 2H; CH₂), 3.9 (s,3H; CH₃), 4.2 (multi, H; CH-NH₂), 4.8 (d, H, J=4 Hz; CH-OH), 7.1, 7.3(d, d, 2H, 2H, J=9 Hz; ##STR18##

(6) Preparation of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoicacid

(2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid (2.70 g, 12.0mmol), 2.52 ml (18.0 mmol) of triethylamine and 3.95 g (14.4 mmol) ofbenzyl S-4,6-dimethylpyrimidin-2-ylthiocarbonate were dissolved in amixed solvent of 12 ml of water and 12 ml of dioxane.

The reaction was completed through stirring overnight at roomtemperature. The reaction solution was concentrated under reducedpressure in order to distil off dioxane. To the residue was added 50 mlof water and the solution was washed with each 50 ml of ethyl acetatetwice. The aqueous phase was adjusted with diluted hydrochloric acid topH 1-2. Deposited oily products were extracted twice each with 50 ml ofethyl acetate. The extracts were combined, washed with each 50 ml of aaqueous solution of sodium chloride for three times and then dried overanhydrous sodium sulfate.

The sodium sulfate was removed by filtration. The filtrate wasconcentrated under reduced pressure and the residue was triturated withpetroleum ether. Separated crystals were collected by filtration, washedwith petroleum ether and dried in vacuo.(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoicacid was obtained. Yield 3.25 g (75.4%). Mp. 160°-162° C. [α]₅₇₈ ²⁷+87.1° (c=1, acetic acid). NMR Spectrum (DMSO-d₆) δ=2.8 (d, 2H, J=6 Hz;CH₂ -CH), 3.7 (s, 3H; CH₃) 4.0 (d, H, J=2 Hz; CH-OH), 4.1 (multi, H;CH-NH), 5.0 (s, 2H; CH₂ -O), 7.1 (multi, 10H ##STR19## and NH).

(7) Preparation of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucinebenzyl ester(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoicacid (1.44 g, 4.00 mmol), 1.91 g (4.80 mmol) of the p-toluenesulfonicacid salt of benzyl (S)-leucinate and 0.65 g (4.8 mmol) of1-hydroxybenzotriazole were dissolved in 23 ml of tetrahydrofuran. Whilecooling in an ice-salt bath, 0.67 ml (4.8 mmol) of triethylamine and0.99 g (4.8 mmol) of dicyclohexylcarbodiimide were added to the solutionand allowed to react overnight.

Separated dicyclohexylurea was filtered off and the filtrate wasconcentrated under reduced pressure. The residue was dissolved in 40 mlof ethyl acetate and insoluble products were separated by filtrationonce more and washed with ethyl acetate. The filtrate and washingsolution were joined and washed with 0.5 N hydrochloric acid twice, anaqueous solution of sodium chloride for three times, 5% aqueous solutionof sodium hydrogen carbonate twice and an aqueous solution of commonsalt for three times successively and then dried over anhydrous sodiumsulfate.

The sodium sulfate was separated out by filtration and the filtrate wasconcentrated under reduced pressure. The residue was triturated withn-hexane and precipitated crystals were collected by filtration, washedwith n-hexane and dried in vacuo. The benzyl ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 2.21 g (98.2%). Mp. 124°-126° C. [α]₅₇₈ ²⁷ +21.6°(c=1, acetic acid). NMR Spectrum (CDCl₃) δ=0.9 (d, 6H, J=5 Hz; (CH₃)₂CH) 2.9 (d, 2H, J=8 Hz; CH--CH₂ --Ar), 3.7 (s, 3H; CH₃ --O), 5.0, 5.1(s, s, 2H, 2H; CH₂ --OCOCH, CH₂ --OCONH) 5.5 (d, H, J=9 Hz; NH), 7.0(multi, 15H; ##STR20## and NH).

(8) Preparation of (2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucine

The benzyl ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucine(1.69 g, 3.00 mmol) was dissolved in 30 ml of 95% acetic acid. Acatalytic amount of palladium black was added and hydrogen wasintroduced under atmospheric pressure to the solution.

When the catalytic reduction was completed at room temperature in 4.5hours, the palladium black was separated by filtration and the filtratewas well concentrated under reduced pressure to dryness. To the residuewas added 20 ml of acetone. Deposited crystals were collected byfiltration and then washed with acetone and dried in vacuo.(2S,3R)-3-amino-2-hydroxy-4-(4-methoxyphenyl)butanoyl-(S)-leucine wasobtained. Yield 0.95 g (94%). Mp. 228°-231° C. (decomposition). [α]₅₇₈³¹ -12.6° (c=1, acetic acid). NMR Spectrum (CF₃ COOD) δ=1.1 (d, 6H, J=5Hz; (CH₃)₂ CH), 3.3 (d, 2H, J=7 Hz; CH₂ --Ar), 4.0 (s, 3H; CH₃ --O), 4.1(multi, H; CH--NH₂), 4.7 (multi, H; CH--NH) 4.8 (d, H, J=4 Hz; CH--OH)7.0, 7.3 (d, d, 2H, 2H, J= 9 Hz; ##STR21##

EXAMPLE 2 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-methoxyphenyl)butanoicacid

N-[2-oxo-2-(2-methoxyphenyl)ethyl]acetamide (m.p. 75°-78° C.) (16.6 g,80.0 mmol) and 37.6 g (0.450 mol) of sodium hydrogen carbonate weredissolved in a mizture of 92.7 ml (0.450 mol) of 25% glyoxylic acidaqueous solution and 150 ml of methanol. When the reaction mixture wasallowed to react at 50°-60° C., the reaction was completed after 40hours.

The reaction solution was treated in the same manner as in Example 1 (1)andthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-methoxyphenyl)butanoicacid was obtained. Yield 13.2 g (58.6%). Mp. 164°-165° C. (foaming) NMRSpectrum (DMSO-d₆) δ=1.9 (s, 3H ; CH₃ --CO), 3.9 (s, 3H; CH₃ --O), 4.5(d, H, J=3 Hz; CH--OH), 5.8 (d, d, H; CH--NH), 7.5 (multi, 5H; ##STR22##and NH).

(2) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(2-methoxyphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(2-methoxyphenyl)butanoicacid (1.41 g, 5.00 mmol) was dissolved in a mixture of 30 ml of aceticacid and 5 ml of methanol. When 0.3 g of 10% palladium carbon was addedand hydrogen was passed at 60° C. under atmospheric pressure, thereaction was completed after about 7.5 hours.

The reaction solution was treated in the same manner as in Example 1(2)and threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-methoxyphenyl)butanoic acidwas obtained. Yield 1.13 g (84.5%). Mp. 195°-198° C. (decomposition) NMRSpectrum (CF₃ COOD)δ=2.3 (s, 3H; CH₃ --CO), 3.2 (d, 2H, J=8 Hz; CH₂),4.0 (s, 3H; CH₃ --O), 4.6 (d, H, J=2 Hz; CH--OH), 4.9 (multi, H;CH--NH), 7.1 (multi, 4H; ##STR23##

(3) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoicacid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-methoxyphenyl)butanoic acid(0.50 g, 1.9 mmol) was dissolved in a mixture of 3 ml of 2 Nhydrochloric acid and 3 ml of dioxane. When the reaction mixture washeated at 50° C., reaction was completed after 20 hours. The reactionsolution was adjusted with 5 N sodium hydroxide aqueous solution to pH7. To the above solution were added 0.35 ml (3.7 mmol) of triethylamineand 0.77 g (2.8 mmol) of benzylS-4,6-dimethylpyrimidin-2-iythiocarbonate in 2 ml of dioxane.

The reaction was completed through stirring overnight at roomtemperature. The reaction solution was treated in the same manner as inExample 1(6). Semi-solidthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoicwas obtained. Yield 0.42 g (63%).

(4) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucinebenzyl ester

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoicacid (0.42 g, 1.2 mmol), 0.56 g (1.4 mmol) of the p-toluenesulfonic acidsalt of (S)-leucine benzyl ester and 0.19 g (1.4 mmol) of1-hydroxybenzotriazole were dissolved in 13 ml of tetrahydrofuran. Whilecooling in an ice-salt bath, 0.20 ml (1.4 mmol) of triethylamine and0.24 g (1.2 mmol) of dicyclohexylcarbodiimide were added and thereaction mixture was allowed to react overnight. The reaction solutionwas treated in the same manner as in Example 1(7). The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 0.39 g (59%). Mp. 90°-92° C. NMR Spectrum (CDCl₃)δ=0.9 (d, 6H, J=4 Hz; (CH₃)₂ CH), 3.0 (d, 2H, J=8 Hz; CH--CH₂ --Ar), 3.8(s, 3H; CH₃ --O), 5.0, 5.1 (s, s, 2H, 2H; CH₂ --OCONH, CH₂ --OCOCH), 5.3(d, H, J=9 Hz; NH), 7.0 (multi, 15H; ##STR24## and NH).

(5) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucine

The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucine(0.34 g, 0.60 mmol) was dissolved in 5 ml of acetic acid, to which acatalytic amount of palladium black was added and then hydrogen wasintroduced under atmospheric pressure. When the reaction mixture wasallowed to react at room temperature, the catalytic reduction wascompleted after 7 hours.

The reaction solution was treated in the same manner as in Example 1(8).Threo-(2RS)-3-amino-2-hydroxy-4-(2-methoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 118 mg (58%). Mp. 212°-215° C. (decomposition).[α]₅₇₈ ²³ -12.2° (c=0.5, acetic acid). NMR Spectrum (CF₃ COOD) δ=1.1(dd, 6H; (CH₃)₂ CH), 3.3 (d, 2H, J=7 Hz; CH₂ --Ar), 3.9 (s, 3H; CH₃--O), 4.2 (multi, H; CH--NH₂), 4.7 (d, H, J=3 Hz; CH--OH), 4.7 (multi,H; CH--NH), 7.0 (multi, 4H ##STR25## .

EXAMPLE 3 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-methoxyphenyl)butanoicacid

N-[2-oxo-2-(3-methoxyphenyl)ethyl]acetamide (m.p. 104°-107° C.) (3.00 g,14.5 mmol) and 10.2 g (0.121 mol) of sodium hydrogen carbonate weredissolved in a mixture of 20.4 ml (78.2 mmol) of 25% glyoxylic acidaqueous solution and 30 ml of methanol. When the reaction mixture wasallowed to react at 50°-60° C., the reaction was completed overnight.The reaction solution was concentrated under reduced pressure and theresidue was dissolved in water and then washed with ethyl acetate. Theaqueous phase was separated and adjusted with hydrochloric acid to pH1-2. Deposited oily products were extracted with ethyl acetate for threetimes. The extracts were combined, washed with an aqueous solution ofsodium chloride and then dried over anhydrous sodium sulfate.

The sodium sulfate was separated out by filtration, and the filtrate wasconcentrated to dryness under reduced pressure. To the residue was addeda small amount of ethyl acetate and precipitated crystals were collectedby filtration, washed with ethyl acetate and then dried in vacuo.Three-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-methoxyphenyl)butanoicacid was obtained. Yield 3.51 g (85.2%). Mp. 159°-162° C.(decomposition). NMR Spectrum (DMSO-d₆) δ=2.0 (s, 3H; CH₃ -CO), 3.9 (s,3H; CH₃ -O), 4.5 (d, H, J=4 Hz; CH-OH), 5.7 (dd, H; CH-NH), 7.4 (multi,4H; ##STR26## 8.2 (d, H, J=9 Hz; NH).

(2) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(3-methoxyphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(3-methoxyphenyl)butanoicacid (1.41 g, 5.00 mmol) was dissolved in 50 ml of acetic acid, to which1.4 g of 10% palladium carbon was added and hydrogen was introduced at60° C. under atmospheric pressure. The reaction was completed afterabout 5 hours.

The reaction solution was treated in the same manner as in Example 1(2).Threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-methoxyphenyl)butanoic acid wasobtained. Yield 0.81 g (61%). Mp. 169°-172° C. (decomposition). NMRSpectrum (CF₃ COOD) δ=2.4 (s, 3H; CH₃ -CO), 3.2 (d, 2H, J=8 Hz; CH₂),4.1 (s, 3H; CH₃ -O), 4.7 (d, H, J=2 Hz; CH-OH), 4.9 (multi, H; CH-NH),7.2 (multi, 4H; ##STR27##

(3) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoicacid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-(3-methoxyphenyl)butanoic acid(0.70 g, 2.6 mmol) was dissolved in a mixture of 3 ml of 2N hydrochloricacid and 3 ml of dioxane. When the reaction mixture was allowed to reactat 50° C., the reaction was completed after 20 hours.

The reaction solution was adjusted with 5N sodium hydroxide aqueoussolution to pH 7. To the above solution were added 0.50 ml (5.2 mmol) oftriethylamine and 1.08 g (3.93 mmol) of benzylS-4,6-dimethylpyrimidin-2-ylthiocarbonate in 3 ml of dioxane.

When the reaction mixture was allowed to react under stirring at roomtemperature, the reaction was completed overnight. The reaction solutionwas treated in the same manner as in Example 1(6). Semi-solidthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoicacid was obtained. Yield 0.63 g (67%).

(4) Preparation of benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)-leucine

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoicacid (0.63 g, 1.8 mmol), 0.84 g (2.1 mmol) of the p-toluenesulfonic acidsalt of (S)-leucine benzyl ester and 0.28 g (2.1 mmol) of1-hydroxybenzotriazole were dissolved in 20 ml of tetrahydrofuran. Whilecooling with a mixture of common salt and ice, 0.30 ml (2.1 mmol) oftriethylamine and 0.36 g (1.8 mmol) of dicyclohexylcarbodiimide wereadded and the reaction mixture was allowed to react overnight. Thereaction solution was treated in the same manner as in Example 1(7). Thebenzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)leucinewas obtained. Yield 0.52 g (53%). Mp. 61°-65° C. NMR Spectrum (CDCl₃)δ=0.9 (d, 6H, J=4 Hz; (CH₃)₂ CH), 2.9 (d, 2H, J=7 Hz; ##STR28## 3.7 (s,3H; CH₃ -O), 5.0, 5.1 (s, s, 2H, 2H; CH₂ -OCONH, CH₂ -OCOCH), 5.5 (d, H,J=9 Hz; NH), 7.0 (multi, 15H; ##STR29## and NH).

(5) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)-leucine

The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)-leucine(0.45 g, 0.80 mmol) was dissolved in 7 ml of acetic acid, to which acatalytic amount of palladium black was added and hydrogen wasintroduced under atmospheric pressure. The catalytic hydrogenation wascompleted at room temperature after 8 hours.

The catalyst was separated out by filtration and the filtrate was wellconcentrated under reduced pressure to dryness. To the residue was added10 ml of acetone. Precipitated crystals were collected by filtration anddissolved in 1N hydrochloric acid. Insoluble materials were filtratedoff with the addition of a small amount of activated carbon and thefiltrate was adjusted with aqueous ammonia to pH 5-6. Separated crystalswere collected by filtration, washed with water and then acetone anddried in vacuo.Threo-(2RS)-3-amino-2-hydroxy-4-(3-methoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 137 mg (51%). Mp. 210°-213° C. (decomposition).[α]₅₇₈ ²³ - 9.5° (c=0.5, acetic acid). NMR Spectrum (CF₃ COOD) δ=1.0(dd, 6H; (CH₃)₂ CH), 3.2 (multi, 2H; CH₂ -Ar), 4.0 (s, 3H; CH₃ -O), 4.2(multi, H; CH-NH₂), 4.7 (multi, H; CH-NH) 4.7 (d, H, J=3 Hz; CH-0H), 7.2(multi, 4H; ##STR30##

EXAMPLE 4 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-phenylphenyl)butanoicacid

N-[2-oxo-2-(4-phenylphenyl)ethyl]acetamide (m.p 144°-146° C.) (19.8 g,78.0 mmol), 18.4 g (0.218 mol) of sodium hydrogen carbonate and 14.4 g(0.156 mol) of glyoxylic acid monohydrate were dissolved in a mixedsolvent of 10 ml of water and 150 ml of methanol. When the solution wasallowed to react at 50°-60° C., the reaction was completed overnight.The reaction solution was treated in the same manner as in Example 1(1).Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-phenylphenyl)butanoicacid was obtained. Yield 25.8 g (100%). Mp. 159°-161° C.(decomposition). NMR Spectrum (CF₃ COOD) δ=2.2 (s, 3H; CH₃), 5.1 (d, H,J=3 Hz; CH-OH), 6.2 (dd, H; CH-NH), 7.8 (multi, 9H; ##STR31##

(2) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-phenylphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-phenylphenyl)butanoicacid (4.91 g, 15.0 mmol) was dissolved in a mixed solvent of 135 ml ofacetic acid and 30 ml of methanol and 1.0 g of 10% palladium carbon wasadded to the solution. When the mixture was allowed to react in the flowof hydrogen at 60° C. under atmospheric pressure, the reaction wascompleted after about 11 hours. The reaction mixture was treated in thesame manner as in Example 1(2).Threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-phenylphenyl)butanoic acid wasobtained. Yield 3.62 g (77.0%). Mp. 175°-176° C. NMR Spectrum (CF₃ DOOD)δ=2.3 (s, 3H; CH₃), 3.1 (2H; CH₂), 4.8 (multi, 2H, CH-NH, CH-OH) 7.5(multi, 9H; ##STR32##

(3) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(4-phenylphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-(4-phenylphenyl)butanoic acid(3.13 g, 10.0 mmol) was dissolved in a mixed solvent of 12 ml of 2Nhydrochloric acid and 12 ml of dioxane. When the solution was allowed toreact heated at 60° C., the reaction was completed after 24 hours. Thereaction solution was decolorized by the addition of small amount ofactivated carbon. After separating out the activated carbon byfiltration, the filtrate was adjusted with concentrated aqueous ammoniato pH 5-6. After ice cooling, deposited crystals were collected byfiltration, washed with cold acetone and then dried in vacuo.Threo-(2RS)-3-amino-2-hydroxy-4-(4-phenylphenyl)butanoic acid wasobtained. Yield 2.23 g (82.2%). Mp. 253°-257° C. (decomposition). NMRSpectrum (CF₃ COOD) δ=3.2 (2H; CH₂), 4.3 (multi, H; CH-NH₂), 4.9 (d, H,J=16 Hz; CH-OH), 7.5 multi, 9H; ##STR33##

(4) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoicacid

Threo-(2RS)-3-amino-2-hydroxy-4-(4-phenylphenyl)butanoic acid (1.90 g,7.00 mmol), 1.96 ml (14.0 mmol) of triethylamine and 2.88 g (10.5 mmol)of benzyl S-4,6-dimethylprimidin-2-ylthiolcarbonate were dissolved in amixed solvent of 7 ml of water and 10 ml of dioxane.

When the solution was allowed to react under stirring at roomtemperature, the reaction was completed overnight. The reaction solutionwas concentrated under reduced pressure. The concentrated solution wasincorporated with water and then adjusted with hydrochloric acid to pH1 - 2. Precipitated crystals were collected by filration, washed withwater and then with ethyl acetate successively and dried in vacuo.Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoicacid was obtained. Yield 1.76 g (62.0%). Mp. 179°-181° C. NMR Spectrum(DMSO-d₆) δ=2.8 (broad, 2H; CH₂ -CH), 4.1 (multi, 2H; CH-OH, CH-NH), 5.0(s, 2H; CH₂ -O), 7.3 (multi, 15H; ##STR34## and NH).

(5) Preparation of benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoyl-(S)-leucine

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoicacid (1.42 g, 3.50 mmol), 1.67 g (4.20 mmol) of the p-toluenesulfonicacid salt of benzyl (S)-leucinate and 0.57 g (4.2 mmol) of1-hydroxybenzotriazole were dissolved in 40 ml of tetrahydrofuran. Whilecooling with common salt and ice, were added 0.50 ml (4.2 mmol) oftriethylamine and 0.72 g (3.5 mmol) of dicyclohexylcarbodiimide and theywere allowed to react overnight.

The reaction mixture was treated in the same manner as in Example 1(7).The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoyl-(S)-leucinewas obtained. Yield 1.35 g (63.4%). Mp. 146°-149° C. NMR Spectrum(CDCl₃) δ=1.0 (d, 6H, J=4 Hz; (CH₃)₂ CH), 3.1 (d, 2H, J=7 Hz; CH-CH₂-Ar), 5.1, 5.3 (d, s, 2H, 2H, J=2 Hz; CH₂ -OCOCH, CH₂ -OCONH), 5.5 (d,H, J=9 Hz; NH), 7.5 (multi, 20H; ##STR35## and NH).

(6) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(4-phenylphenyl)butanoyl-(S)-leucine

The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-phenylphenyl)butanoyl-(S)-leucine(1.22 g, 2.00 mmol) was dissolved in a mixed solvent of 15 ml of aceticacid, 15 ml of ethyl acetate and 10 ml of methanol. A catalytic amountof palladium black was added to the solution and hydrogen was introducedunder atmospheric pressure. The catalytic hydrogenation was completed atroom temperature after about 8 hours. The reaction mixture was treatedin the same manner as in Example 1(8).Threo-(2RS)-3-amino-2-hydroxy-4-(4-phenylphenyl)butanoyl-(S)-leucine wasobtained. Yield 0.75 g (98%). Mp. 218°-221° C. (decomposition). [α]₅₇₈²⁶ - 12.9° (c=0.5, acetic acid). NMR Spectrum (CF₃ COOD) δ=1.1 (d, 6H,J=4 Hz; (CH₃)₂ CH), 3.2 (d, 2H, J=7 Hz; CH₂ -Ar), 4.2 (multi, H,CH-NH₂), 4.8 (multi, H; CH-NH), 5.0 (d, H, J=4 Hz; CH-OH), 7.5 (multi,9H; ##STR36##

EXAMPLE 5 (1) Preparation ofthreo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-(4-isopropylphenyl)butanoicacid

N-[2-oxo-2-(4-isopropylphenyl)ethyl]benzamide (m.p. 115°-117° C.) (3.38g, 12.0 mmol), 2.82 g (33.6 mmol) of sodium hydrogen carbonate and 2.21g (24.0 mmol) of glyoxylic acid monohydrate were dissolved in a mixedsolvent of 20 ml of methanol, 10 ml of ethanol and 2 ml of water. Whenthe solution was allowed to react at 50°-60° C., the reaction wascompleted overnight. The reaction solution was concentrated underreduced pressure to dryness. The residue was dissolved in water andwashed with ethyl acetate. The aqueous phase was separated and adjustedwith hydrochloric acid to pH 1 - 2. Deposited oily product was extractedwith ethyl acetate twice. The extracts were joined, washed with waterand then dried over anhydrous sodium sulfate. The sodium sulfate wasseparated out by filtration and the filtrate was concentrated underreduced pressure. The residue was crystallized from ether andisopropylether.Threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(4-isopropylphenyl)butanoicacid was obtained. Yield 2.27 g (53.2%). Mp. 153°-154° C. NMR Spectrum(DMSO-d₆) δ=1.3 (d, 6H, J=7 Hz; (CH₃)₂ CH), 3.0 (multi, H; (CH₃)₂ CH),4.7 (d, H, J=4 Hz; CH-OH), 6.0 (dd, H; CH-NH), 7.7 (multi, 9H; ##STR37##8.4 (d, H, J=9 Hz; NH).

(2) Preparation ofthreo-(2RS)-3-benzoylamino-2-hydroxy-4-(4-isopropylphenyl)butanoic acid

Threo-(2-RS)-3-benzoylamino-2-hydroxy-4-oxo-4-(4-isopropylphenyl)butanoicacid (1.78 g, 5.00 mmol) was dissolved in 82 ml of acetic acid. When thesolution was hydrogenated with addition of 0.89 g of 10% palladiumcarbon while introducing hydrogen at 60° C. under atmospheric pressure,the reaction was completed after about 6 hours. The catalyst wasseparated by filtration and the filtrate was concentrated under reducedpressure to dryness. The residue was crystallized fromchloroform-petroleum ether.Threo-(2-RS)-3-benzoylamino-2-hydroxy-4-(4-isopropylphenyl)butanoic acidwas obtained. Yield 1.46 g (85.5%). Mp. 91°-94° C. NMR Spectrum(DMSO-d₆) δ=1.3 (d, 6H, J=7 Hz; (CH₃)₂ CH), 3.0 (d, 2H, J=7 Hz; CH₂),4.1 (d, H, J=3 Hz; CH-OH), 4.6 (multi, H; CH-NH), 7.7 (multi, 10H;##STR38## and NH).

(3) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(4-isopropylphenyl)butanoic acid

Threo-(2RS)-3-benzoylamino-2-hydroxy-4-(4-isopropylphenyl)butanoic acid(1.20 g, 3.53 mmol) was dissolved in a mixed solvent of 1 ml of 10Nhydrochloric acid and 1 ml of dioxane. When the solution was allowed toreact under reflux, the reaction was completed after 6 hours. Water wasadded to the reaction solution which was then washed with ethyl acetate.The aqueous phase was separated and adjusted with 5N sodium hydroxideaqueous solution to pH 5 - 6. Deposited crystals were collected byfiltration, washed with cold water and dried in vacuo.Threo-(2RS)-3-amino-2-hydroxy-4-(4-isopropylphenyl)butanoic acid wasobtained. Yield 0.50 g (60%).

(4) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoicacid

Threo-(2RS)-3-amino-2-hydroxy-4-(4-isopropylphenyl)butanoic acid (0.50g, 2.10 mmol), 0.44 ml (3.2 mmol) of triethylamine and 0.69 g (2.5 mmol)of benzyl S-4,6-dimethylpyrimidin-2-ylthiolcarbonate were dissolved in amixed solvent of 2.5 ml of water and 2.5 ml of dioxane. When thesolution was allowed to react under stirring at room temperature, thereaction was completed overnight. The reaction solution was treated inthe same manner as in Example 1(6).Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoicacid was obtained. Yield0.43 g (55%). Mp. 129°-130° C. NMR Spectrum(CDCl₃) δ=1.3 (d, 6H, J=7 Hz; (CH₃)₂ CH), 2.9 (d, 2H, J=7 Hz; CH₂ -CH),4.2 (d, H, J=2 Hz; CH-OH), 4.3 (multi, H; CH-NH), 5.1 (s, 2H; CH₂ -O),7.2 (multi, 10H; ##STR39## and NH).

(5) Preparation of benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoyl-(S)-leucine

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoicacid (0.37 g, 1.0 mmol), 0.48 g (1.2 mmol) of the p-toluenesulfonic acidsalt of benzyl (S)-leucinate and 0.16 g (1.2 mmol) of1-hydroxybenzotriazole were dissolved in 12 ml of tetrahydrofuran. Whilecooling in an ice-common salt bath, 0.17 ml (1.2 mmol) of triethylamineand 0.25 g (1.2 mmol) of dicyclohexylcarbodiimide were added. Thesolution was allowed to react overnight. The reaction solution wastreated in the same manner as in Example 1(7). The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoyl-(S)-leucinewas obtained. Yield 0.51 g (88%). Mp. 107°-110° C. NMR Spectrum (CDCl₃)δ=0.9 (d, 6H, J=5 Hz; (CH₃)₂ CH-CH₂), 1.3 (d, 6H, J=7 Hz; (CH₃)₂ CH-Ar),5.1, 5.2 (s, s, 2H, 2H; CH₂ -OCOCH, CH₂ -OCONH), 5.5 (d, H, J=9 Hz; NH),7.1 (s, 4H; ##STR40## 7.3 (s, 10H; ##STR41##

(6) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(4-isopropylphenyl)butanoyl-(S)-leucine

The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropylphenyl)butanoyl-(S)-leucine(0.29 g, 0.50 mmol) was dissolved in 11 ml of 90% acetic acid. Acatalytic amount of palladium black was added to the solution andhydrogen was introduced under atmospheric pressure. The catalyticreduction was completed at room temperature after 7 hours. The reactionmixture was treated in the same manner as in Example 1(8).Threo-(2RS)-3-amino-2-hydroxy-4-(4-isopropylphenyl)butanoyl-(S)-leucinewas obtained. Yield 141 mg (80%). Mp. 234°-237° C. (decomposition).[α]₅₇₈ ²⁹ -5.9° (c=0.5, acetic acid). NMR Spectrum (CF₃ COOD) δ=1.1 (d,6H, J=6 Hz; (CH₃)₂ CH-CH₂), 1.3 (d, 6H, J=7 Hz; (CH₃)₂ CH-Ar), 3.0(multi, 3H; CH₂ -Ar, CH-Ar), 4.2 (multi, H; CH-NH₂) 4.8 (d, multi, 2 H,J=3 Hz; CH-NH, CH-OH), 7.3 (multi, 4H; ##STR42##

EXAMPLE 6 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-isopropoxyphenyl)butanoicacid

N-[2-oxo-2-(4-isopropoxyphenyl)ethyl]acetamide (m.p.. 92°-95° C.) 1.90g, 8.05 mmol), 1.89 g (22.5 mmol) of sodium hydrogen carbonate and 1.33(14.5 mmol) of glyoxylic acid monohydrate were dissolved in 30 ml ofmethanol. When the solution was allowed to react at 50°-60° C., thereaction was completed after 6.5 hours. The reaction solution wasconcentrated under reduced pressure to dryness and the residue wasdissolved in 30 ml of water. Insoluble compounds were separated out withethyl acetate and the aqueous phase was adjusted with dilutedhydrochloric acid to pH 1 - 2. Deposited oily product was extracted with30 ml of ethyl acetate twice. The extracts were combined and dried overanhydrous sodium sulfate. The sodium sulfate was separated by filtrationand the filtrate was concentrated under reduced pressure. The residuewas triturated with 20 ml of ether and precipitated crystals werecollected by filtration, washed with ether and dried in vacuo.Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-isopropoxyphenyl)butanoic acid wasobtained. Yield 1.47 g (58.8%). Mp. 157°-158° C. (decomposition). NMRSpectrum (DMSO-d₆) δ=1.3 (d, 6H, J=6 Hz; (CH₃)₂ CH), 1.8 (s, 3H; CH₃CO), 4.4 (d, H, J=4 Hz; CH-OH), 4.7 (multi, H; (CH₃)₂ CH), 5.9 (dd, H;CH-NH), 7.2, 8.0 (d, d, 2H, 2H, J=9 Hz; ##STR43##

(2) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-isopropoxyphenyl)butanoicacid (1.50 g, 4.80 mmol) was dissolved in 20 ml of methanol, to which0.2 g of 10% palladium carbon was added. When the solution wascatalytically reduced in an autoclave under a hydrogen pressure of 20Kg/cm² at 40° C. for 1 hour and then at 90° C. for 4 hours, the reactionwas completed. The catalyst was separated out by filtration and thefiltrate was concentrated to dryness under reduced pressure. The residuewas dissolved in 20 ml of ethyl acetate and then dicyclohexylamine wasadded so as to adjust pH to 7, whereby crystals were deposited.Precipitated crystals were collected by filtration, washed with ethylacetate and dried in vacuo. The dicyclohexylamine salt ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoic acidwas obtained. Yield 1.90 g (81.3%). Mp. 182°-184° C. The above acid,after removed dicyclohexylamine in the same manner as described in thesucceeding Example 6(3), showed the following NMR spectrum. NMR Spectrum(DMSO-d₆), δ=1.2 (d, 6H, J=6 Hz; (CH₃)₂ CH), 1.8 (s, 3H; CH₃ -CO), 2.7(d, 2H, J=8 Hz; CH₂), 3.9 (d, H, J=3 Hz; CH-OH), 4.5 (multi, H; CH-NH),6.8, 7.2 (d, d, 2H, 2H, J=8 Hz; ##STR44## 7.6 (d, H, J=8 Hz; NH).

(3) Preparation ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoicacid

The dicyclohexylamine salt ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoic acid(1.9 g, 3.90 mmol) was added to a mixed solvent of 20 ml of N sulfuricacid and 20 ml of ethyl acetate and shaken. The ethyl acetate phase wasseparated, washed with water and then concentrated under reducedpressure to dryness. The residue was dissolved in a mixture of 10 ml ofconcentrated hydrochloric acid and 10 ml of dioxane. When the solutionwas heated at 70° C., the reaction was completed after 1 hour.

The reaction solution was concentrated under reduced pressure todryness. The residue was dissolved in a mixture of 10 ml of water and 10ml of dioxane. After adjusting pH to 8 - 9 by the addition oftriethylamine, 1.60 g (5.90 mmol) of benzylS-4,6-dimethylpyrimidin-2-ylthiolcarbonate was further added. When thesolution was allowed to react while stirred at room temperature, thereaction was completed after two days. The reaction solution was treatedin the same manner as in Example 1(6).Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoicacid was obtained. Yield 0.41 g (26%). Mp. 135°-137° C. NMR Spectrum(DMSO-d₆) δ=1.2 (d, 6H, J=6 Hz; (CH₃)₂ CH), 2.7 (d, 2H, J=8 Hz; CH₂-CH), 4.0 (d, H, J=2 Hz; CH-OH), 4.5 (multi, H; CH-NH), 5.0 (s, 2H, CH₂-O), 7.0 (multi, 10H; ##STR45## and NH).

(4) Preparation of benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoyl-(S)-luecine

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoicacid (0.40 g, 1.0 mmol), 0.44 g (1.1 mmol) of the p-toluenesulfonic acidsalt of benzyl (S)-leucinate and 0.16 g (1.2 mmol) of1-hydroxybenzotriazole were dissolved in 5 ml of tetrahydrofuran, towhich 0.13 ml (1.1 mmol) of triethylamine and 0.25 g (1.2 mmol) ofdicyclohexylcarbodiimide were added, while cooling with common salt andice. The solution was allowed to react overnight. The reaction solutionwas treated in the same manner as in Example 1(7). The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 0.24 g (40%). Mp. 120°-125° C. NMR Spectrum (CDCl₃)δ=0.9 (d, 6H, J=6 Hz; (CH₃)₂ CH-CH₂), 1.2 (d, 6H, J=6 Hz; (CH₃)₂ CH-O),2.9 (d, 2H, J= 6 Hz; CH-CH₂ -Ar), 4.1 (d, H, J=2 Hz; CH-OH), 4.5 (multi,H; CH-NH), 5.0, 5.1 (s, s, 2H, 2H; CH₂ -O-COCH, CH₂ -OCONH), 5.5 (d, H,J=8 Hz; NH), 7.1 (multi, 15H; ##STR46## and NH).

(5) Preparation ofthreo-(2RS)-3-amino-2-hydroxy-4-(4-isopropoxyphenyl)butanoyl-(S)-leucine

The benzyl ester ofthreo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(4-isopropoxyphenyl)butanoyl-(S)-leucine(0.20 g, 0.33 mmol) was dissolved in 9 ml of 90% acetic acid. Acatalytic amount of palladium black was added to the solution andhydrogen was introduced at atmospheric pressure. The catalytic reductionwas completed at room temperature after 7 hours. The reaction solutionwas treated in the same manner as in Example 1(8).Threo-(2RS)-3-amino-2-hydroxy-4-(4-isopropoxyphenyl)butanoyl-(S)-leucinewas obtained. Yield 74 mg (59%). Mp. 210°-215° C. (decomposition). NMRSpectrum (CF₃ COOD) δ=1.0 (d, 6H, J=5 Hz; (CH₃)₂ CH-CH₂), 1.4 (d, 6H,J=6Hz; (CH₃)₂ CH-O), 3.2 (d, 2H, J=6Hz; CH-CH₂ -Ar), 4.2 (multi, H;(CH₃)₂ CH-O), 4.7 (multi, 3H; CH-OH, CH-NH₂, CH-NH), 7.1, 7.3 (d, d, 2H,J=10Hz; ##STR47##

EXAMPLE 7 (1) Preparation of methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl(butanoicacid

N-[2-oxo-2-(4-methoxyphenyl)ethyl]acetamide (4.14 g, 20.0 mmol), 3.52 g(40.0 mmol) of the methyl ester of glyoxylic acid and 2.52 g (30.0 mmol)of sodium hydrogen carbonate were dissolved in a mixed solvent of 50 mlof methanol and 3 ml of water. When the solution was allowed to react at50°-60° C., the reaction was completed overnight. The reaction solutionwas concentrated to dryness under reduced pressure. The residue wasdissolved in 50 ml of ethyl acetate, and washed with water, a dilutedaqueous solution of sodium hydrogen sulfite and then water successively.The ethyl acetate phase was separated and dried over anhydrous sodiumsulfate. The sodium sulfate was separated out by filtration and thefiltrate was concentrated under reduced pressure. Oily product obtainedwas subjected to silica gel column chromatography usingchloroform-methanol (9:1 v/v) as a solvent. Fractions containing theaimed product were collected and concentrated under reduced pressure.The methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid was obtained. Yield 2.67 g (45.3%). Mp. 148°-151° C. NMR SpectrumDMSO-d₆) δ=1.9 (s, 3H; CH₃ -CO), 3.6, 3.9 (s, s, 3H, 3H; CH₃ -OCO, CH₃-OAr), 4.5 (d, H, J=4 Hz; CH-OH), 5.7 (dd, H; CH-NH), 7.1, 7.9 (d, d,2H, 2H, J=9 Hz ##STR48## 8.2 (d, H, J=9Hz; NH).

(2) Preparation of methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid

The methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-methoxyphenyl)butanoicacid (2.00 g, 6.78 mmol) was dissolved in 50 ml of methanol, to which0.20 g of 10% palladium carbon was added. When the soution was reducedin an autoclave under a hydrogen pressure of 30 kg/cm² at 40° C. for 1hour and then at 80°-90° C. for 5 hours, the reaction was completed. Thecatalyst was separated out by filtration and the filtrate wasconcentrated to dryness under reduced pressure. Crystals deposited fromether were collected by filtration, washed with ether and dried invacuo. The methyl ester ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic acid wasobtained. Yield 1.62 g (85.0%). Mp. 105°-110° C. NMR Spectrum (DMSO-d₆)δ-1.8 (s, 3H; CH₃ -CO), 2.8 (d, 2H, J=7Hz; CH₂), 3.6, 3.7 (s, s, 3H, 3H;CH₃ -OCO, CH₃ -OAr), 4.1 (d, H, J=2Hz; CH-OH), 4.3 (multi, H; CH-NH),6.8, 7.2 (d, d, 2H, 2H, J=9Hz; ##STR49## 7.4 (d, H, J=9Hz; NH).

Other compounds listed in Table 2 were also prepared in analogousprocedures to those in the foregoing examples. As the starting compoundsof formula (III), those corresponding compounds represented by theformula (I) or (II) were employed. All of the compounds are ofthreo-(2RS) form unless otherwise referred to for their optical isomericform.

                                      TABLE 2                                     __________________________________________________________________________    Compounds and physical properties thereof                                      Examples No.                                                                             ##STR50##                                                                                               ##STR51##                               __________________________________________________________________________    8          R.sub.1 = 4-isobutylphenyl, R.sub.2 = benzoylamino                                                      R.sub.1 = 4-isobutylphenyl, R.sub.2                                           = benzoylamino                                      R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 155-158° C. (decomposition)                                                                 m.p. 127-130° C.                             NMR Spectrum (DMSOd.sub.6)                                                                              NMR Spectrum (DMSOd.sub.6) δ =                                          0.9(d,6H,                                           δ = 0.9(d,6H,J = 7Hz; (CH.sub.3).sub.2 CH),                                                       J = 7Hz; (CH.sub.3).sub.2 CH), 1.8                                            (multi, H;                                          1.9(multi, H; (CH.sub.3).sub.2 CH), 2.6(d,2H,J                                                          (CH.sub.3).sub.2 CH), 2.4(d,2H,J =                                            7Hz; (CH.sub.3).sub.2 CHCH.sub.2)                   CH.sub.2), 4.7(d,H,J = 3Hz; CHOH),                                                                      3.0(d,H,J = 8Hz; CH.sub.2 CHCH),                     ##STR52##                                                                                               ##STR53##                               __________________________________________________________________________    9          R.sub.1 = 4-isobutoxyphenyl, R.sub.2 = acetylamino                                                      R.sub.1 = 4-isobutoxyphenyl, R.sub.2                                          = acetylamino                                       R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 125-127° C. (foaming)                                                                       m.p. 135-140° C.                             NMR Spectrum (DMSOd.sub.6)                                                    δ = 1.0(d,6H,J = 7Hz; (CH.sub.3).sub.2 CH),                             1.9(s,3H; CH.sub.3CO), 3.8(d,2H,J = 6Hz; CH.sub.2),                           4.4(d,H,J = 4Hz; CHOH), 5.8(dd,H; CHNH),                                       ##STR54##                                                                    8.0(d,H,J = 8Hz; NH)                                               __________________________________________________________________________    10         R.sub.1 =  4-fluorophenyl, R.sub.2 = acetylamino                                                        R.sub.1 = 4-fluorophenyl, R.sub.2 =                                           acetylamino                                         R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 152-154° C.   m.p. 190-191° C.                             NMR Spectrum (DMSOd.sub.6) δ = 1.9(s,3H;                                                          NMR Spectrum (CF.sub.3 COOD) δ                                          = 2.3(s,3H; CH.sub.3),                              4.4(d,H,J = 4Hz; CHOH), 5.6(dd,H; CHNH)                                                                 3.1(d,2H,J = 8Hz; CH.sub.2),                                                  4.6(d,H,J = 3Hz;                                     ##STR55##                                                                                               ##STR56##                               __________________________________________________________________________    11         R.sub.1 = 3-hydroxyphenyl, R.sub.2 = acetylamino,                                                       R.sub.1 = 3-hydroxyphenyl, R.sub.2 =                                          acetylamino,                                        R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 184-187° C. (decomposition)                                                                 m.p. 182-191° C.                                                       (decomposition)                                     NMR Spectrum (DMSOd.sub.6) δ = 1.9(s,3H;                                                          NMR Spectrum (CF.sub.3 COOD) δ                                          = 2.3(s,3H; CH.sub.3),                              4.5(d,H,J = 3Hz; CHOH), 5.7(dd,H; CHNH)                                                                 3.1(d,2H,J = 7Hz; CH.sub.2),                                                  4.7(d,H,J = 2Hz;                                     ##STR57##                                                                                               ##STR58##                               __________________________________________________________________________    12         R.sub.1 = 1-naphthyl, R.sub.2 = acetylamino,                                                            R.sub.1 = 1-naphthyl, R.sub.2 =                                               acetylamino,                                        R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 162-163° C. (decomposition)                                                                 m.p. 164-168° C.                             NMR Spectrum (DMSOd.sub.6), δ = 2.0(s,3H;                                                         NMR Spectrum (DMSOd.sub.6), δ                                           = 1.8(s,3H; CH.sub.3),                              4.5(d,H,J = 4Hz; CHOH), 5.8(dd,H; CHNH)                                                                 3.2(d,2H,J = 7Hz; CH.sub.2)                          ##STR59##                                                                                               ##STR60##                               __________________________________________________________________________    13         R.sub.1 = 2-naphthyl, R.sub.2 = acetylamino,                                                            R.sub.1 = 2-naphthyl, R.sub.2 =                                               acetylamino,                                        R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               m.p. 138.5-141.5° C. (decomposition)                                                             m.p. 174-178° C.                             NMR Spectrum (DMSOd.sub.6)                                                                              NMR Spectrum (CF.sub.3 COOD) δ                                          = 2.2(s,3H; CH.sub.3)                               δ = 2.0(s,3H; CH.sub.3), 4.6(d,H,J = 4Hz;                                                         3.2(d,2H,J = 8Hz; CH.sub.2),                                                  4.6(d,H,J = 3Hz;                                     ##STR61##                                                                                               ##STR62##                               __________________________________________________________________________    14         R.sub.1 = 2-methoxyphenyl, R.sub.2 = acetylamino                                                        R.sub.1 = 2-methoxyphenyl, R.sub.2 =                                          acetylamino                                         R.sub.3 = hydrogen        R.sub.3 = hydrogen                       __________________________________________________________________________               See Example 2-(1)         See Example 2-(2)                        __________________________________________________________________________    15         R.sub.1 = 3,4-dihydroxyphenyl, R.sub.2 = acetylamino                                                    R.sub.1 = 3,4-dihydroxyphenyl,                      R.sub.3 = hydrogen        R.sub.2 = acetylamino, R.sub.3 =                                              hydrogen                                 __________________________________________________________________________               m.p. 146-150° C. (decomposition)                                                                 m.p. 235-239° C.                                                       (decomposition)                                     NMR Spectrum (DMSOd.sub.6) δ = 1.9(s,3H;                                                          NMR Spectrum (CF.sub.3 COOD) δ                                          = 2.3(s; CH.sub.3),                                 4.4(d,H,J = 4Hz; CHOH), 5.6(dd,H; CHNH),                                                                4.7, 4.8(d, multi, H,H; CHOH,                                                 CHNH),                                               ##STR63##                                                                                               ##STR64##                                          7.9(d,H,J = 9Hz; NH).                                              __________________________________________________________________________    Compounds and physical properties thereof                                      Examples No.                                                                         ##STR65##                                                             __________________________________________________________________________    8      R.sub.1 = 4-isobutylphenyl, R'.sub.2 = amino                                                              R.sub. 1 = 4-isobutylphenyl, R'.sub.2                                         = benzyloxylcarbonylamino                  __________________________________________________________________________           m.p. 240-245° C. (decomposition)                                                                   m.p. 155-160° C.                                                       NMR Spectrum (DMSOd.sub.6)                                                    δ = 0.9(d,6H,J = 7Hz;                                                   (CH.sub.3).sub.2 CH),                                                         1.8(multi, H; (CH.sub.3).sub.2 CH),                                           2.4(d,2H,J = 7Hz; (CH.sub.3).sub.2                                            CHCH.sub.2),                                                                  2.8(d,2H,J = 6Hz; CH.sub.2 CHCH),                                             4.1(multi, 2H;                                                                CHOH, CHNH), 4.9(d,2H,J = 3Hz;                                                CH.sub.2),                                                                     ##STR66##                                 __________________________________________________________________________    9                                  R.sub.1 = 4-isobutoxyphenyl,                                                  R'.sub.2 = benzyloxycarbonylamino          __________________________________________________________________________                                       m.p. 172-180° C.                    __________________________________________________________________________    10     R.sub.1 = 4-fluorophenyl,                                                     R'.sub.2 = benzyloxycarbonylamino                                      __________________________________________________________________________           m.p. 126-132° C.                                                       NMR Spectrum (DMSOd.sub.6)                                                    δ = 2.8(d,2H,J = 6Hz; CH.sub.2CH),                                      4.0(multi, 2H; CHNH, CHOH),                                                    ##STR67##                                                                     ##STR68##                                                             __________________________________________________________________________    11     R.sub.1 = 3-hydroxyphenyl,                                                    R'.sub.2 = benzyloxycarbonylamino                                      __________________________________________________________________________           m.p. 120-125° C.                                                       NMR Spectrum (DMSOd.sub.6)                                                    δ = 2.7(d,2H,J = 7Hz; CH.sub.2CH),                                      4.0(d,H,J = 3Hz; CHOH),                                                       4.1(multi, H; CHNH), 5.0(s,2H; CH.sub.2O),                                     ##STR69##                                                             __________________________________________________________________________    12     R.sub.1 = 1-naphthyl, R'.sub.2 = amino                                                                    R.sub.1 = 1-naphthyl, R'.sub.2 =                                              benzyloxycarbonylamino                     __________________________________________________________________________           m.p. 214-215° C. (decomposition)                                                                   m.p. 110-120° C.                           NMR Spectrum (CF.sub.3 COOD)                                                                              NMR Spectrum (CDCl.sub.3)                         δ = 3.5(d,2H,J = 8Hz; CH.sub.2),                                                                    δ = 2.8(d,2H,J = 8Hz;                                                   CH.sub.2CH)                                       4.2(multi, H; CHNH.sub.2),  4.1(d,H,J = 2Hz; CHOH),                            ##STR70##                                                                                                 ##STR71##                                 __________________________________________________________________________    13     R.sub.1 = 2-naphthyl, R'.sub.2 = amino                                                                    R.sub.1 = 2-naphthyl, R'.sub.2 =                                              benzyloxycarbonylamino                     __________________________________________________________________________                                       m.p. 172-174° C.                           NMR Spectrum (CF.sub.3 COOD)                                                                              NMR Spectrum (DMSOd.sub.6)                        δ = 3.4(dd,2H; CH.sub.2), 4.3(multi, H; CHNH),                                                      δ = 3.0(d,2H,J = 8Hz;                                                   CH.sub.2CH),                                      4.7(d,H,J = 3Hz; CHOH),     4.0(d,H,J = 3Hz; CHOH),                            ##STR72##                                                                                                 ##STR73##                                 __________________________________________________________________________    14     R.sub.1 = 2-hydroxyphenyl, R'.sub.2 = amino                                                               R.sub.1  = 2-hydroxyphenyl,                       (*See Note 1)               R'.sub.2 = benzyloxycarbonylamino          __________________________________________________________________________           m.p. 235-239° C.     m.p. 110-115° C.                           NMR Spectrum (CF.sub.3 COOD)                                                                              NMR Spectrum (CDCl.sub.3)                         δ = 3.4(d,2H,J = 7Hz; CH.sub.2), 4.3(multi,                                                         δ= 2.9(d,2H,J = 7Hz;                                                    CH.sub.2CH),                                      CHNH), 4.7(d,H,J = 4Hz; CHOH),                                                                            4.2(d,H,J = 2Hz; CHOH), 4.3(multi, H;              ##STR74##                                                                                                 ##STR75##                                 __________________________________________________________________________    15     R.sub.1 = 3,4-dihydroxyphenyl, R'.sub.2 = benzyloxycarbonylamino       __________________________________________________________________________           A NMR Spectrum (DMSOd.sub.6) δ = 2.8(d,2H,J = 8Hz;                        CH.sub.2CH),                                                                  4.1, 4.2(d, multi, H,H,J = 2Hz; CHOH, CHNH), 5.0(s,2H;                        CH.sub.2O),                                                                    ##STR76##                                                                  B m.p. 129-133° C.                                                        ##STR77##                                                                    *See Note 2                                                          __________________________________________________________________________        The compound of the formula (VII')                                         No.plesExam-                                                                      ##STR78##                                                                                                   ##STR79##                                  __________________________________________________________________________    8   R.sub.1 = 4-isobutylphenyl, R".sub.2 = benzyloxycarbonyl-                                                   R.sub.1 = 4-isobutylphenyl, R.sub.4 =                                         isobutyl                                        amino, R.sub.4 = isobutyl, P = benzyloxycarbonyl                          __________________________________________________________________________        m.p. 110-113° C.       m.p. 221- 226° C.,                                                     [a].sup.29 .sub.578 - 10.1° (c =                                       0.5, acetic acid)                               NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD)                    δ = 0.9(d,6H,J = 7Hz; (CH.sub.3).sub.2 CHCH.sub.2Ar),                                                 δ = 0.9(d,6H,J = 7Hz;                                                   (CH.sub.3).sub.2 CHCH.sub.2),                   0.9(d,6H,J = 7Hz; (CH.sub.3).sub.2 CHCH.sub.2 CH),                                                          1.1(d,6H,J = 7Hz; (CH.sub.3).sub.2                                            CHCH.sub.2),                                    2.4(d,2H,J = 7Hz; (CH.sub.3).sub.2 CHCH.sub.2Ar),                                                           2.5(d,2H,J = 7Hz; (CH.sub.3).sub.2                                            CHCH.sub.2Ar),                                  2.9(multi, 2H; CHCHCH.sub.2Ar),                                                                             3.2(multi, 2H; CHCHCH.sub.2Ar),                 5.0, 5.1(s,s,2H,2H; CH.sub.2OCOCH,                                                                          4.2(multi, H; CHNH.sub.2), 4.8(multi,                                         H;                                              CH.sub.2OCONH), 5.3(d,H,J = 9Hz; NH),                                                                       CHNH), 5.0(d,H,J = 4Hz; CHOH),                   ##STR80##                                                                                                   ##STR81##                                  __________________________________________________________________________    9   R.sub.1 = 4-isobutoxyphenyl, R".sub. 2 = benzyloxycarbonyl-                                                 R.sub.1 = 4-isobutylophenyl, R.sub.4 =                                        isobutyl                                        amino, R.sub.4 = isobutyl, P = benzyloxycarbonyl                          __________________________________________________________________________        m.p. 131-136° C.       m.p. 210-215° C.                     __________________________________________________________________________    10  R.sub.1 = 4-fluorophenyl,     R.sub.1 = 4-fluorophenyl, R.sub.4 =                                           isobutyl                                        R".sub.2 = benzyloxycarbonylamino,                                            R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 94-101° C.        m.p. 220-225° C.                                                       (decomposition),                                                              [a].sup.27 .sub.578 - 4.0° (c =                                        0.9, acetic acid)                               NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD) δ =                                        1.0(dd,6H; (CH.sub.3).sub.2 CH),                δ = 0.8(d,6H; (CH.sub.3).sub.2 CH), 2.9(d,2H,7Hz;                                                     3.2(d,2H,J = 6Hz; CH.sub.2Ar),                                                4.2(multi, H; CHNH.sub.2),                       ##STR82##                                                                                                   ##STR83##                                  __________________________________________________________________________    11  R.sub.1 = 3-hydroxyphenyl,    R.sub.1 = 3-hydroxyphenyl, R.sub.4 =                                          isobutyl                                        R".sub.2 = benzyloxycarbonylamino,                                            R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 141-145° C.       NMR Spectrum (CF.sub.3 COOD)                    NMR Spectrum (CDCl.sub.3)     δ = 1.1(dd,6H; (CH.sub.3).sub.2                                         CH),                                            δ = 0.8(d,6H,J = 5Hz; (CH.sub.3).sub.2 CH),                                                           3.3(d,2H,J = 6Hz; CH.sub.2Ar),                                                4.2(multi, H; CHNH.sub.2),                       ##STR84##                                                                                                   ##STR85##                                  __________________________________________________________________________    12  R.sub.1 = 1-naphthyl,         R.sub.1 = 1-naphthyl, R.sub.4 =                                               isobutyl                                        R".sub.2 = benzyloxycarbonylamino                                             R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 124-128° C.       m.p. 185-191° C.                                                       (decomposition)                                 NMR Spectrum (CDCl.sub.3)                                                     δ = 0.9(d,6H,J = 4Hz; (CH.sub.3).sub.2 CH),                              ##STR86##                                                                    2H; CH.sub.2OCONH, CH.sub.2OCOCH),                                             ##STR87##                                                                     ##STR88##                                                                __________________________________________________________________________    13  R.sub.1 = 2-naphthyl,         R.sub.1 = 2-naphthyl, R.sub.4 =                                               isobutyl                                        R".sub.2 = benzyloxycarbonylamino,                                            R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 119-122° C.       m.p. 226-229° C.                                                       (decomposition)                                                               [a] .sup.25 .sub.578 - 6.5° (c =                                       1, acetic acid).                                NMR Spectrum (CDCl.sub.3) δ = 0.9(dd,6H; (CH.sub.3)CH),                                               NMR Spectrum (CF.sub.3 COOD) δ =                                        1.0(dd,6H;                                       ##STR89##                                                                                                   ##STR90##                                       ##STR91##                                                                                                   ##STR92##                                       ##STR93##                                                                __________________________________________________________________________    14  R.sub.1 = 2-hydroxyphenyl,    R.sub.1 = 2-hydroxyphenyl, R.sub.4 =                                          isobutyl                                        R".sub.2 = benzyloxycarbonylamino                                             R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 120-130° C.       m.p. 238-244° C.                                                       (decomposition)                                                               [a].sup.29 .sub.578 - 7.7° (c =                                        0.5, acetic acid)                               NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD) δ =                                        1.1(dd,6H;                                      δ = 0.9(d,6H,J = 5Hz; (CH.sub.3).sub.2 CH)                                                            (CH.sub.3).sub.2 CH), 3.2(d,2H,J = 7Hz;                                       CH.sub.2Ar),                                    3.0(d,2H,J = 8Hz; CHCH.sub.2Ar),                                                                            4.3(multi, H; CHNH.sub.2), 4.7(multi,                                         2H;                                             5.0, 5.1(s,s,2H,2H; CH.sub.2OCOCH, CH.sub.2OCONH),                                                          CHOH, CHNH),                                     ##STR94##                                                                                                   ##STR95##                                  __________________________________________________________________________    15  R.sub.1 = 3,4-dihydroxyphenyl,                                                                              R.sub.1 = 3,4-dihydroxyphenyl, R.sub.4                                        = isobutyl                                      R".sub.2 = benzyloxycarbonylamino,                                            R.sub.4 = isobutyl, P = benzyloxycarbonyl                                 __________________________________________________________________________        m.p. 70-75° C.         m.p. 175-185° C.                                                       (decomposition)                                                               [a] .sup.29 .sub.578 - 13.3° (c                                        = 0.5, acetic acid)                             NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD) δ =                                        1.1(dd,6H;                                      δ = 0.8(d,6H; J = 4Hz; (CH.sub.3).sub.2 CH).                                                          (CH.sub.3).sub.2 CH), 3.2(d,H,J = 6Hz;                                        CH.sub.2Ar),                                                                  4.2(multi, H; CHNH.sub.2), 4.9(multi,                                         2H;                                                                            ##STR96##                                  __________________________________________________________________________    16  R.sub.1 = 4-methoxyphenyl,    R.sub.1 = 4-methoxyphenyl,                      R".sub.2 = benzyloxycarbonylamino                                                                           R.sub.4 = isopropyl                             R.sub.4 = isopropyl, P = benzyloxycarbonyl,                                                                 (2S,3R)-Form                                    (2S,3R)-Form                                                              __________________________________________________________________________        m.p. 123-125° C.       m.p. 240-243° C.                                                       (decomposition)                                 [a].sup.28 .sub.578 + 22.8° (c = 1, acetic acid)                                                     [a].sup.31 .sub.578 + 5.0° (c =                                        1, acetic acid)                                 NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD)                    δ = 0.8(dd,6H; (CH.sub.3).sub.2 CH),                                                                  δ = 1.1(d,6H,J = 7Hz;                                                   (CH.sub.3).sub.2 CH),                           2.1(multi, H; (CH.sub.3).sub.2 CH), 2.9(d,2H,J = 7Hz;                                                       2.4(multi, H; (CH.sub.3).sub.2 CH)              CHCH.sub.2Ar), 3.9(s,3H; CH.sub.3O),                                                                        3.3(multi, 2H; CH.sub.2Ar), 4.0(s,3H;                                         CH.sub.3O)                                      4.2(dd, multi, H,H; CHOH, CH.sub.2CHNH)                                                                     4.1(multi, H; CHNH.sub.2),                      4.6(dd,H; COCHNH),            4.6(d,H,J = 6Hz; CHNH), 4.9(d,H,J =                                           3Hz;                                            5.0, 5.1(s,s,2H,2H; CH.sub.2OCOCH, CH.sub.2OCONH),                                                          CHOH), 7.1, 7.3(d,d,2H,2H,J = 9Hz;               ##STR97##                                                                                                   ##STR98##                                  __________________________________________________________________________    17  R.sub.1 = 2-naphthyl,         R.sub.1 = 2-naphthyl                            R".sub.2 = benzyloxycarbonylamino,                                                                          R.sub.4 = 1-methylpropyl                        R.sub.4 = 1-methylpropyl, P = benzyloxycarbonyl                           __________________________________________________________________________        m.p. 106-111° C.       m.p. 225-230°  C.                                                      (decomposition)                                                               [a].sup.31 .sub.578 + 17.3° (c =                                       1, acetic acid)                                 NMR Spectrum (CDCl.sub.3)     NMR Spectrum (CF.sub.3 COOD)                    δ = 0.8, 0.9(d,t,3H,3H,J = 6Hz; J = 6Hz; CH.sub.3CH,                                                  δ = 1.1(d,t,3H,3H; CH.sub.3CH,                                          CH.sub.3CH.sub.2),                              CH.sub.3CH.sub.2), 3.1(d,2H,J = 7Hz; CHCH.sub.2Ar),                                                         3.4(d,2H,J = 6Hz; CH.sub.2Ar),                  4.2(d,H,J = 2Hz; CHOH),       4.3(multi, H; CHNH.sub.2)                       4.6(multi, H; CHNH),          4.7(d,H,J = 5Hz; CHNH),                         4.9, 5.1(s,s,2H,2H; CH.sub.2OCOCH, CH.sub.2OCONH),                                                          4.9(d,H,J = 4Hz; CHOH),                          ##STR99##                                                                                                   ##STR100##                                 __________________________________________________________________________     Note 1:                                                                       Preparation of threo(2RS)-3-amino-2-hydroxy-4-(2-hydroxyphenyl)butanoic       acid                                                                          Threo-(2RS)-3-acetylamino-2-hydroxy-4-(2-methoxyphenyl)butanoic acid (2.6     g, 10.0 mmol) prepared in Example 2(2) was added to 12 ml of 47%              hydrobromic acid.                                                             When the mixture was heated at 100° C., the reaction was completed     after 6 hours. The reaction mixture was concentrated to dryness under         reduced pressure. The                                                         residue was dissolved in 10 ml of water and concentrated under reduced        pressure to dryness. The above dissolving and concentrating procedure was     repeated once                                                                 again Finally, the residue was dissolved in 10 ml of water and the aqueou     phase was adjusted with concentrated aqueous ammonia to pH 5-6. After ice     cooling,                                                                      deposited crystals were collected by filtration, washed with cold water       and dried in vacuo.                                                           Threo(2RS)-3-amino-2-hydroxy-4-(2-hydroxyphenyl)butanoic acid was             obtained (1.40 g).                                                            Note 2:                                                                       Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(3,4-dihydroxyphenyl)but    noic acid prepared by reaction of                                              threo-(2RS)-3-acetylamino-2-hydroxy-4-(3,4-dihydroxyphenyl)butanoic acid      and benzyl S--4,6dimethylpyrimidin-2-ylthiolcarbonate was isolated in the     same manner as                                                                the isolation of the reaction product in Example 1(6). The isolated           compound was hydroscopic. NMR spectrums of the isolated compound are show     in the column A                                                               of the Table.                                                                 A portion of the crystals was dissolved in acetone and the solution was       neutralized by the addition of dicyclohexylamine. Deposited crystals were     collected by filtration,                                                      washed with acetone and dried in vacuo. Melting point and NMR spectrums o     the dicyclohexylamine salt prepared are shown in the column B of the          Table.                                                                        Note 3:                                                                       The starting material used here was                                           (2S,3R)3-benzyloxycarbonylamino-2-hydroxy-4-(4-methoxyphenyl)butanoic aci     prepared in Example 1(6).                                                     Note 4:                                                                       The starting material used here was                                           threo(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-(2-naphthyl)butanoic acid     the compound of the formula (V),                                              prepared in Example 12.                                                       Note 5:                                                                       The physical properties of the following compounds, which is used as          materials in Examples 8 and 10, are as follows;                               N--[2-oxo-2-(4-isobutylphenyl)ethyl]benzamide: m.p 106-109° C.         N--[2-oxo-2-(4-fluorophenyl)ethyl]acetamide: m.p 153-155° C.      

EXAMPLE 18 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-bromophenyl)butanoic acid

N-[2-oxo-2-(4-bromophenyl)ethyl]acetamide (m.p. 173°-175° C.) (17.0 g,66.0 mmol), 15.8 g (0.188 mol) of sodium hydrogen carbonate and 10.9 g(0.120 mol) of glyoxylic acid monohydrate were dissolved in the a mixedsolvent of 170 ml of methanol and 20 ml of water. When the solution wasallowed to react at 50°-60° C., the reaction was completed after 5hours. The reaction solution was treated in the same manner as inExample 1(1).Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-(4-bromophenyl)butanoic acidwas obtained. Yield 12.7 g (58.4%). Mp. 177°-180° C. (decomposition).NMR Spectrum (DMSO-d₆) δ-1.8 (s, 3H; CH₃), 4.5 (d, H, J=5Hz; CH-OH), 5.6(dd, H; CH-NH), 7.7 (multi, 4H; ##STR101## 8.1 (d, H, J=8Hz; NH).

EXAMPLE 19 (1) Preparation ofthreo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid

N-(2-oxo-2phenylethyl)acetamide (4.43 g, 0.025 mol) and 4.20 g (0.05mol) of sodium hydrogen carbonate were dissolved in a mixed solvent of13.0 g (0.044 mol) of 25% glyoxylic acid aqueous solution and 25 ml ofwater. When the solution was allowed to react at 50°-60° C., thereaction was completed overnight. The reaction solution was cooled withice and adjusted with diluted hydrochloric acid to pH 1-2. Depositedcrystals were collected by filtration, washed with water and dried invacuo over phosphorous pentoxide.Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid wasobtained. Yield 4.07 g (64.9%). Mp. 151°-152° C. (decomposition). NMRSpectrum (DMSO-d₆) δ=2.0 (s, 3H; CH₃) 4.6 (d, H, J=3 Hz; CH-OH), 5.9(dd, H; CH-NH) 7.7 (multi, 5H; ##STR102## In addition broad absorptionderived from NH and OH was shown at 6.6-8.0, which was eliminated by theaddition of deuterated water.

(2) Preparation of threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoicacid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid (3.00 g,0.012 mol) was dissolved in 25 ml of acetic acid. When the solution washydrogenated with the addition of 0.30 g of 5% palladium carbon andintroduction of hydrogen under atmospheric pressure at 60° C., thereaction was completed after about 6 hours.

After separating out the catalyst by filtration, the filtrate wasconcentrated under reduced pressure. The residue was triturated with 20ml of ethyl acetate. Deposited crystals were collected by filtration,washed with ethyl acetate and dried in vacuo.Threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid was obtained.Yield 2.33 g (82.3%). Mp. 174°-176° C.

In addition, broad absorption derived from OH was shown at 7.0-8.0,which disappeared by the addition of deuterated water.

(3) Preparation of (2S,3R)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid

Threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid (10.87 g,0.046 mol) and 5.55 g (0.046 mol) of S(-)-1-phenylethylamine wasdissolved under heating in 90 ml of ethanol and then the solution wasallowed to cool at room temperature. Deposited crystals were collectedby filtration, washed with a small amount of ethanol and dried in vacuo.6.37 g of optically impure crystals were obtained. [α]_(D) ²⁰ +16.8°(c=1, methanol)

The crystals (6.30 g) were dissolved under heating in 100 ml of ethanoland then allowed to cool to room temperature. Deposited crystals werecollected by filtration, washed with a small amount of ethanol and driedin vacuo. The S(-)-1-phenylamine salt of(2S,3R)-3-acetylamino-2-hydroxy-phenylbutanoic acid was obtained. Yield3.45 g. Mp. 194°-195° C. [α]_(D) ²⁰ +29.0° (c=1, methanol)

Specific optical rotation of the authetic salt prepared from(2S,3R)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid andS(-)-1-phenylethylamine was as follows: [α]_(D) ²⁰ +29.1° (c=1,methanol).

(4) Preparation of (2S,3R)-3-amino-2-hydroxy-4-phenylbutanoic acid

The S(-)-1-phenylethylamine salt of(2S,3R)-3-acetylamino-2-hydroxy-4-phenylbutanoic acid (4.25 g, 0.0119mol) and 1.49 g (0.0178 mol) of sodium hydrogen carbonate were dissolvedin 80 ml of water, from which S(-)-1-phenylethylamine was extracted forthree times each with 50 ml of ethyl acetate.

The aqueous phase was adjusted with concentrated hydrochloric acid to pH1-2, and concentrated under reduced pressure to about 40 ml. When theconcentrated solution was refluxed under heating with addition of 1.7 ml(0.02 mol) of concentrated hydrochloric acid, the reaction was completedafter two hours.

The reaction solution was concentrated to dryness under reducedpressure. The residue was dissolved in 10 ml of water and thenconcentrated to dryness. The above dissolving and concentratingprocedure was repeated.

Finally, the residue was dissolved in 40 ml of water and the solutionwas adjusted with an aqueous solution of 2 N sodium hydroxide to pH 5-6.After ice cooling, deposited crystals were collected by filtration,washed with cold water and dried in vacuo.(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoic acid was obtained. Yield 1.48g (63.8%). [α]₅₇₈ ¹⁹.5 +32.5° (c=0.76, N HCl). The reference value ofthe compound (J. Med. Chem., 20, 510 (1977)), [α]₅₇₈ ²⁰˜25 +29.5° (c=1,N HCl)

(5) Preparation of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid

(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoic acid (1.45 g, 7.44 mmol),1.13 g (11.2 mmol) of triethylamine and 2.24 g (8.20 mmol) of benzylS-4,6-dimethylpyrimidin-2-ylthiolcarbonate were dissolved in a mixedsolvent of 7 ml of water and 7 ml of dioxane. When the solution wasallowed to react under stirring at room temperature, the reaction wascompleted after 3 hours.

To the reaction solution was added 20 ml of water and the solution waswashed twice each time with 25 ml of ethyl acetate. The aqueous phasewas adjusted with diluted hydrochloric acid to pH 1-2. Deposited oilyproduct was extracted twice each time with 30 ml of ethyl acetate. Theextracts were joined, washed for three times each time with 30 ml of acommon salt aqueous solution and dried over anhydrous sodium sulfate.

After separating out the sodium sulfate by filtration, the filtrate wasconcentrated to dryness under reduced pressure. The residue wastriturated with petroleum ether. Precipitated crystals were collected byfiltration, washed with petroleum ether and dried in vacuo.(2S,3R)-3-benzyl-oxycarbonylamino- 2-hydroxy-4-phenylbutanoic acid wasobtained. Yield 2.10 g (85.7%). Mp. 154°-155° C. [α]₅₇₈ ²⁰ +82.5° (c=1,acetic acid). The reference values of the compound (Example 1(1) inJapanese Patent Unexamined Publication No. 136118/1977), Mp. 154.5° C.[α]₅₇₈ ²⁴ +83.5° (c=1.34, acetic acid).

(6) Preparation of the benzyl ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoyl-(S)-leucine

(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid (2.00g, 6.00 mmol), 2.63 g (6.60 mmol) of the p-toluenesulfonic acid salt ofbenzyl (S)-leucinate and 0.97 g (7.20 mmol) of 1-hydroxybenzotriazolewere dissolved in 20 ml of tetrahydrofuran. While cooling with commonsalt and ice, 0.67 g (6.6 mmol) of triethylamine and 1.49 g (7.20 mmol)of dicyclohexylcarbodiimide were added and the solution was allowed toreact overnight.

Deposited dicyclohexylurea was separated out by filtration and thefiltrate was concentrated to dryness under reduced pressure. To theresidue was added 50 ml of ethyl acetate. After separating Insolubleproducts were separated out by filtration and washed with a small amountof ethyl acetate. The filtrate and the washing solution were joined andwashed twice with 0.5 N hydrochloric acid, three times with a commonsalt aqueous solution, twice with 5% sodium hydrogen carbonate aqueoussolution and then three times with a common salt aqueous solutionsuccessively, and dried over anhydrous sodium sulfate.

After separating the sodium sulfate by filtration, the filtrate wsconcentrated to dryness under reduced pressure and the residue wastriturated with n-hexane. Separated crystals were collected byfiltration, washed with n-hexane and dried in vacuo. The benzyl ester of(2S,3R)-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoyl-(S)-leucinewas obtained. Yield 3.19 g (99.4%). Mp. 122°-123° C. [α]₅₇₈ ²⁴ +15.2°(c=1, acetic acid). The reference values of the compound (Example 2(3)in Japanese Patent Unexamined Publication No. 136118/1977), Mp. 122° C.[α]₅₇₈ ²³ +15.1° (c=0.77, acetic acid).

(7) Preparation ofbestatin[(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-leucine]

The benzyl ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoyl-(S)-leucine(3.00 g, 5.60 mmol) was dissolved in 50 ml of 95% acetic acid. When acatalytic amount of palladium black was added to the solution andhydrogen was introduced under atmospheric pressure, the catalyticreduction was completed after two hours. After separating palladiumblack by filtration, the filtrate was thoroughly concentrated to drynessunder reduced pressure and the residue was triturated with 30 ml ofacetone. Deposited crystals were separated by filtration and dissolvedin 1 N hydrochloric acid. After separating insoluble materials by theaddition of a small amount of activated carbon, the filtrate wasadjusted with diluted aqueous ammonia to pH 5-6. Precipitated crystalswere collected by filtration, washed with acetone and dried in vacuo.Bestatin[(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-leucine] wasobtained. Yield 1.47 g (85.0%). [α]₅₇₈ ²⁵ -21.1° (c=1, acetic acid). Thereference value of the compound (Example 2(4) in Japanese PatentUnexamined Publication No. 136118/1977), [α]₅₇₈ ²⁵ -21.8° (c=0.45,acetic acid).

EXAMPLE 20 (1) Preparation of N-hydroxysuccinimied ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid

(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid (6.58g, 0.02 mol) prepared in the same manner as in Example 19(5) and 2.30 g(0.02 mol) of N-hydroxysuccinimide were dissolved in a mixed solvent of60 ml of dioxane and 60 ml of ethyl acetate. After cooling the solutionbelow 0° C., 4.12 g (0.02 mol) of dicyclohexylcarbodiimide was added andthe reaction mixture was allowed to react at the same temperature for 1hour and then at room temperature overnight.

After separating deposited insoluble compounds by filtration, thefiltrate was concentrated under reduced pressure. The residue wassolidified with petroleum ether (bp. 30°-70° C. fraction) andre-precipitated from ethyl acetate and petroleum ether.N-hydroxysuccinimide ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid wasobtained. Yield 6.51 g (76.4%). Mp. 111°-112° C. [α]₅₇₈ ³⁰ +35.4°(c=1.5, acetic acid).

(2) Preparation of(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-arginine hydrochloride

The N-hydroxysuccinimide ester of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid (4.26g, 0.01 mol) was dissolved in 30 ml of dioxane, to which was added asolution of 2.11 g (0.01 mol) of arginine hydrochloride and 1.40 ml(0.01 mol) of triethylamine in 30 ml of water. The solution was allowedto react at room temperature for 2 days.

After the reaction, the solvent was distilled off under reduced pressureand the residue was dissolved in a solution of chloroform and methanol(1:1 v/v). The solution was subjected to column chromatography usingsilica gel H type 60 to collect the fractions containing the aimedproduct and the solvent was distilled off under reduced pressure. Oily(2S,3R)-benzyloxycarbonylamino-b 2-hydroxy-4-phenylbutanoyl-(S)-argininehydrochloride was obtained. Yield 2.02 g (38.7%).

The hydrochloride prepared was dissolved in a mixture of 40 ml ofmethanol and 20 ml of water. The solution was catalytically reduced atroom temperature under atmospheric pressure for 6 hours using palladiumblack.

After separating out the catalyst by filtration, the filtrate wasconcentrated under reduced pressure. Acetone was added to the residue toprecipitate crystals. The deposited crystals were collected byfiltration, washed with acetone and dried in vacuo.(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-(S)-arginine hydrochloridewas obtained. Yield 1.19 g (30.7%). [α]₅₇₈ ³⁰ -6.4° (c=1.2, acetic acid.

Rf value 0.08 (Measured on silica gel 60 F₂₅₄ plate (manufactured byMerck) using as a developing solvent n-BuOH:AcOH:H₂ O=4:1:1).

EXAMPLE 21: Preparation ofthreo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid

N-(2-oxo-2-phenylethyl)benzamide (16.7 g, 0.07 mol) and 13.0 g (0.155mol) of sodium hydrogen carbonate was dissolved in a mixed solvent of3.75 g (0.13 mol) of 25% glyoxylic acid aqueous solution, 100 ml ofwater and 250 ml of methanol. The solution was allowed to react at50°-60° C. overnight.

After separating insoluble products in the reaction solution byfiltration, the filtrate was concentrated under reduced pressure todistil off methanol. Diluted hydrochloric acid was added to theconcentrated solution to adjust pH 1-2. Deposited crystals werecollected by filtration, washed with water and dried in vacuo overphosphorous pentoxide. Crude crystals (17.6 g) were obtained and theywere recrystallized with ethyl acetate.Threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid wasobtained. Yield 13.5 g (61.8%). Mp. 174°-176° C. (decomposition). NMRSpectrum (DMSO-d₆) δ=4.6 (d, H, J=4 Hz; CH-OH), 5.9 (dd, H; CH-NH) 7.7(multi, 10 H; ##STR103## 8.5 (d, H, J=9 Hz; NH).

(2) Preparation of threo-(2RS)-3-benzoylamino-2-hydroxy-4-phenylbutanoic acid

Threo-(2RS)-3-benzoylamino-2-hydroxy-4-oxo-4-phenylbutanoic acid (5.00g, 0.016 mol) and 0.50 g of 10% palladium carbon were added in 90 ml ofacetic acid. When the solution was allowed to react while introducinghydrogen at 70° C. under atmospheric pressure, the reaction wascompleted after about 8 hours.

After separating out the catalyst by filtration, the filtrate wasconcentrated under reduced pressure. The oily residue was thoroughlytriturated with petroleum ether (bp. 30°-70° C. fraction) and thesupernatant liquid was removed by decantation. Upon adding new petroleumether and rubbing the glass wall with a glass rod under cooling,crystallization occured. After left to stand at room temperature for 1hour, the crystals were collected by filtration, washed with petroleumether and dried in vacuo.Threo-(2RS)-3-benzoylamino-2-hydroxy-4-phenylbutanoic acid was obtained.Yield 4.61 g (96.4%). Mp. 144°-145° C. NMR Spectrum (DMSO-d₆) δ=2.9 (d,2 H, J=7 Hz; CH₂), 4.0 (d, H, J=3 Hz; CH-OH), 4.55 (multi, H; CH-NH),7.25, 7.5 (s, milti, 10 H; ##STR104## 7.95 (d, H, J=8 Hz; NH).

(3) Preparation of (2S,3R)-3-benzoylamino-2-hydroxy-4-phenylbutanoicacid

Threo-(2RS)-3-benzoylamino-2-hydroxy-4-phenylbutanoic acid (6.30 g,0.0211 mol) and 2.57 g (0.211 mol) of S(-)-1-phenylethylamine weredissolved under heating in 18 ml of ethanol and then left overnight atroom temperature.

Deposited crystals were collected by filtration and recrystallized fromethanol. The S(-)-1-phenylethylamine salt of(2S,3R)-3-benzoylamino-2-hydroxy-4-phenylbutanoic acid was obtained.Yield 1.22 g (27.5%). Mp. 147°-148° C. [α]₅₇₈ ²⁹ +70.6° (c=1.02, aceticacid).

Elemental analysis for C₂₅ H₂₈ N₂ O₄ : Found: C: 71.67, H: 6.99, N:6.73; Calculated: C: 71.39, H: 6.72, N: 6.67.

The salt prepared (1.00 g, 2.38 mmol) was added to and shaken with amixture of 20 ml of N sulfuric acid and 50 ml of ethyl acetate. Theethyl acetate phase was separated and washed with water repeatedly tillit became neutral. Then the solution was dried over anhydrous magnesiumsulfate. After separating out the magnesium sulfate by filtration, thefiltrate was concentrated under reduced pressure and the residue wascrystallized from ethyl acetate and petroleum ether (bp. 30°-70° C.fraction). Deposited crystals were collected by filtration, washed withthe same mixed solvent as above and dried in vacuo.(2S,3R)-3-Benzoylamino-2-hydroxy-4-phenylbutanoic acid was obtained.Yield 0.64 g. Mp. 172°-173° C. [α]₅₇₈ ²⁵ +109.5° (c=1.1, acetic acid).

Elemental analysis for C₁₇ H₁₇ NO₄ ; Found: C:68.34, H:5.92, N:4.44;Calculated: C:68.19, H:5.73, N:4.68.

EXAMPLE 22 (1) Preparation ofthreo-(2RS)-2-hydroxy-4-oxo-4-phenyl-3-phthaliminobutanoic acid

N-(2-oxo-2-phenylethyl)phthalimide (10.0 g, 0.0377 mol) and 9.00 g(0.107 mol) of sodium hydrogen carbonate were dissolved in a mixedsolvent of 0.1 g (0.068 mol) of 25% glyoxylic acid aqueous solution and20 ml of ethanol. The solution was allowed to react at 50°-60° C. for 24hours.

The reaction solution was concentrated under reduced pressure to distiloff ethanol. The concentrated solution was incorporated and shaken with100 ml of ethyl acetate and 50 ml of 5% sodium hydrogen carbonateaqueous solution. The aqueous phase was separated and adjusted withdiluted hydrochloric acid to pH 1-2. Deposited crystals were collectedby filtration, washed with water and dried in vacuo over phosphorouspentoxide. Threo-(2RS)-2-hydroxy-4-oxo-4-phenyl-3-phthaliminobanoic acidwas obtained. Yield 1.97 g (15.4%). Mp. 168°-170° C. (decomposition).NMR Spectrum (DMSO-d₆) δ=4.55 (d, H, J=4 Hz; CH--OH), 5.8 (dd, H;CH--NH), 7.8 (multi, 9 H; ##STR105## 8.45 (d, H, J=9 Hz; NH).

(2) Preparation of threo-(2RS)-2-hydroxy-4-phenyl-4-phthaliminobutanoicacid

Threo-(2RS)-2-hydroxy-4-oxo-4-phenyl-3-phthaliminobutanoic acid (1.00 g,2.90 mmol) and 0.10 g of 10% palladium black were added to 20 ml ofacetic acid. The mixture was catalytically reduced while introducinghydrogen at 70° C. under atmospheric pressure for 5.5 hours.

The reaction mixture was concentrated under reduced pressure. Petroleumether (bp. 30°-70° C. fraction) and then a small amount of ethyl acetatewere added to the residue to crystallize oily product. Depositedcrystals were collected by filtration, washed with petroleum ether anddried in vacuo. Threo-(2RS)-2-hydroxy-4-phenyl-3-phthaliminobutanoicacid was obtained. Yield 0.55 g (57%). Mp. 97°-103° C. (foaming). NMRSpectrum (DMSO-d₆) δ=2.9 (d, 2H, J=7 Hz; CH₂), 3.9 (d, H, J=3 Hz;CH--OH), 4.4 (multi, H; CH--NH), 7.3, 7.5 (s, multi, 9H; ##STR106##

Other compounds listed in Table 3 were also produced in the proceduresanalogous to those in Examples 18-22. As the starting material for thecompounds of formula (III), corresponding compounds of the formula (I)or (II) were employed. All of the compounds in Table 3 were ofthreo-(2RS) form.

                                      TABLE 3                                     __________________________________________________________________________    Compounds and physical properties                                             The compounds of the formula (III)                                                                       The compounds of the formula                       __________________________________________________________________________                               (IV)                                               23                                                                              R.sub.1 = phenyl, R.sub.2 = chloroacetylamino                                                          R.sub.1 = phenyl, R.sub.2 = chloroacetylamino        R.sub.3 = hydrogen       R.sub.3 = hydrogen                                 __________________________________________________________________________      m.p. 141-142° C. (decomposition)                                                                m.p. 108-110° C.                              NMR Spectrum (DMSOd.sub.6)                                                                             NMR Spectrum (CDCl.sub.3)                             ##STR107##                                                                                             ##STR108##                                        __________________________________________________________________________    24                                                                              R.sub.1 = phenyl, R.sub.2 = t-butyloxycarbonylamino                                                    R.sub.1 = phenyl, R.sub.2 =                                                   t-butyloxycarbonylamino                              R.sub.3 = hydrogen*Note 1                                                                              R.sub.3 = hydrogen                                 __________________________________________________________________________      m.p. 142-143° C. (decomposition)                                                                NMR Spectrum (CDCl.sub.3)                             ##STR109##                                                                                             ##STR110##                                        __________________________________________________________________________    25                                                                              R.sub.1 = 3-hydroxyphenyl, R.sub.2 = benzoylamino                                                      R.sub.1 = 3-hydroxyphenyl, R.sub.2                                            = benzoylamino                                       R.sub.3 = hydrogen       R.sub.3 = hydrogen                                 __________________________________________________________________________      m.p. 176-177° C. (decomposition)                                                                m.p. 80-90° C. (foaming),                                              162-165° C.                                   NMR Spectrum (DMSOd.sub.6)                                                                             NMR Spectrum (DMSOd.sub.6)                            ##STR111##                                                                                             ##STR112##                                        __________________________________________________________________________    26                                                                              R.sub.1 = 4-hydroxyphenyl, R.sub.2 = acetylamino                                                       R.sub.1 = 4-hydroxyphenyl, R.sub.2                                            = acetylamino                                        R.sub.3 = hydrogen       R.sub.3 = hydrogen                                 __________________________________________________________________________      m.p. 179-181° C. (decomposition)                                                                m.p. 194-197° C. (decomposition)              NMR Spectrum (DMSOd.sub.6)                                                                             NMR Spectrum (DMSOd.sub.6)                            ##STR113##                                                                                             ##STR114##                                        __________________________________________________________________________    27                                                                              R.sub.1 = phenyl, R.sub.2 = chloroacetylamino                                                          R.sub.1 = phenyl, R.sub.2 = chloroacetylamino        R.sub.3 = methyl         R.sub.3 = methyl                                   __________________________________________________________________________      m.p. 119-121° C.  NMR Spectrum (CDCl.sub.3)                             ##STR115##                                                                                             ##STR116##                                                                   IR Spectrum                                                                    ##STR117##                                                                   1740 (broad), 1660 (broad), 1605, 1545                                        and 1525 (broad), 1500, 1440, 1410,                                           1270 and 1220 (broad), 1120, 995, 930,                                        915, 860, 750, 700.                                __________________________________________________________________________     *Note 1:                                                                      2-t-butoxycarbonylaminoacetophenone used as the starting material was         prepared by reacting 2aminoacetophenone hydrochloride with tbutyl             S4,6dimethylpyrimidin-2-ylthiolcarbonate and triethylamine in a mixture o     dioxane and water (1:1).                                                 

EXAMPLE 28

Threo-(2RS)-3-amino-2-hydroxy-4-phenylbutanoic acid (39.00 g, 0.200 mol)prepared from threo-(2RS)-3-acetylamino-2-hydroxy-4-phenylbutanoic acidgiven by Example 19(2) upon hydrolysis with hydrochloric acid in thesame manner as in Example 1(4), 65.80 g (0.240 mol) of benzylS-4,6-dimethylpyridmidin-2-ylthiolcarbonate and 42.0 ml (0.300 mol) oftriethylamine were allowed to react in a mixed solvent of 300 ml ofwater and 300 ml of dioxane and treated in the same manner as in Example1(6). Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoicacid was obtained. Yield 61.30 g (93.2%). Mp. 133°-134° C.

Elemental analysis for C₁₈ H₁₉ NO₅ : Found: C: 65.83, H: 5.77, N: 4.13:Calculation: C: 65.62, H: 5.82, N: 4.26.

Threo-(2RS)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid(5.73 g., 0.0174 mol) prepared and 7.47 g (0.0174 mol) of brucinemonohydrate were dissolved under heating in 100 ml of ethyl acetate andleft at room temperature overnight. Deposited crystals were collected byfiltration and recrystallized with ethyl acetate. The brucine salt of(2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acid wasobtained. Yield 3.58 g (56.9%). Mp. 144° C. [α]₅₇₈ ³¹ +35.9° (c=1.2,acetic acid)

The brucine salt (2.00 g, 2.63 mmol) obtained was dissolved in andshaken with a mixture of 50 ml of N hydrochloric acid and 100 ml ofethyl acetate. The ethyl acetate phase was separated and washed withwater repeatedly till it became neutral. Then the solution was driedover anhydrous magnesium sulfate. After separating out the mangesiumsulfate by filtration, the filtrate was concentrated under reducedpressure. The residue was crystallized from ethyl acetate and petroleumether (bp. 30°-70° C. fraction). Deposited crystals were collected byfiltration, washed with the same mixed solvent as above and dried invacuo. (2S,3R)-3-benzyloxycarbonylamino-2-hydroxy-4-phenylbutanoic acidwas obtained. Yield 0.77 g. Mp. 154°-155° C. [α]₅₇₈ ²⁵ +83.0° (c=1.0,acetic acid). Rf value (Example 1(2) in Japanese Patent UnexaminedPublication No. 136118/1977). Mp. 154.5° C. [α]₅₇₈ ²⁴ +83.5° (c=1.34,acetic acid).

What is claimed is:
 1. Threo-3-protected amino-2-hydroxy-4-oxobutanoicacid or its esters represented by the formula: ##STR118## wherein R₁represents naphthyl or a group of the formula ##STR119## in which R₆ andR₇ represent individually hydrogen, halogen, amino or an amino protectedwith acyl, lower alkyloxycarbonyl or carbamoyl, hydroxy or hydroxyprotected with acyl, lower alkyloxycarbonyl or carbamoyl, lower alkoxy,lower alkyl or phenyl group; R₂ represents an amino protected with acyl,lower alkyloxycarbonyl or carbamoyl; and R₃ represents hydrogen, loweralkyl having 1 to 6 carbon atoms, or benzyl.
 2. A compound according toclaim 1, whereinR₁ is phenyl and R₃ is hydrogen.
 3. A compound accordingto claim 1, whereinR₁ is hydroxyphenyl and R₃ is hydrogen.